bluuebunny/crossref_metadata_2025 huggingface dataset:
Clinical Characterization of Epilepsy of Unknown Cause in Cats
Authors: Wahle A.M. , Brühschwein A. , Matiasek K. , Putschbach K. , Wagner E. , Mueller R.S. , Fischer A.
Published: November 2013

Background

The diagnosis of feline epilepsy of unknown cause (EUC) requires a thorough diagnostic evaluation, otherwise the prevalence ofEUCcould be overestimated.

Hypothesis

FelineEUCis a clinically defined disease entity, which differs from feline hippocampal necrosis by the absence of magnetic resonance imaging (MRI) signal alteration of the hippocampus. The objectives of this study were (1) to evaluate the prevalence ofEUCin a hospital population of cats by applying well‐defined inclusion criteria, and (2) to describe the clinical course ofEUC.

Animals

Eighty‐one cats with recurrent seizures.

Methods

Retrospective study—medical records were reviewed for cats presented for evaluation of recurrent seizures (2005–2010). Inclusion criteria were a defined diagnosis based on laboratory data, and eitherMRIor histopathology. Final outcome was confirmed by telephone interview with the owner. Magnetic resonance images were reviewed to evaluate hippocampal morphology and signal alterations.

Results

Epilepsy of unknown cause was diagnosed in 22% of cats with epilepsy. Physical, neurologic, and laboratory examinations, and either 1.5 TMRIand cerebrospinal fluid analysis or postmortem examination failed to identify an underlying cause. Cats withEUChad a higher survival rate (P < .05) and seizure remission occurred frequently (44.4%).

Conclusion and Clinical Importance

A detailed clinical evaluation and diagnostic imaging withMRIis recommended in any cat with recurrent seizures. The prognosis of cats with normalMRIfindings and a clinical diagnosis ofEUCare good. Standardized imaging guidelines should be established to assess the hippocampus in cats.
Comparison of Thoracic Auscultation, Clinical Score, and Ultrasonography as Indicators of Bovine Respiratory Disease in Preweaned Dairy Calves
Authors: Buczinski S. , Forté G. , Francoz D. , Bélanger A.‐M.
Published: November 2013

Background

The diagnostic tools for bovine respiratory disease diagnosis include clinical inspection, thoracic auscultation, and ultrasonography.

Hypothesis

Thoracic auscultation and clinical examination have limitations in the detection of lung consolidation in dairy calves.

Animals

Prospective cohort of 106 preweaned calves from 13 different dairy herds (10 with a history of active bovine respiratory disease (BRD) in calves and 3 without suspectedBRDproblems).

Methods

Each preweaned calf was clinically inspected using the Wisconsin calf respiratory scoring chart (CRSC) and treatment history was noted. Systematic thoracic auscultation and ultrasonography then were performed, the latter focusing on lung consolidation. Mortality was recorded over a 30‐day period.

Results

A total of 56 of 106 calves had ultrasonographic evidence of lung consolidation. The sensitivity of thoracic auscultation to detect consolidation was 5.9% (range, 0–16.7%). Only 41.1% (23/33) of calves with consolidated lungs had been treated previously by the producers. When addingCRSCand previousBRDtreatment by the producer, sensitivity of detection increased to 71.4% (40/56). The area under the receiver operating characteristics curve was 0.809 (95%CI, 0.721–0.879) for the number of areas within the lungs with consolidation and 0.743 (95%CI, 0.648–0.823) for the maximal depth of consolidation as predictors of death within 1 month after examination. These were not significantly different (P = .06).

Conclusions and Clinical Importance

This study shows that thoracic auscultation is of limited value in diagnosing lung consolidation in calves. Ultrasonographic assessment of the thorax could be a useful tool to assessBRDdetection efficiency on dairy farms.
Genomic inference accurately predicts the timing and severity of a recent bottleneck in a nonmodel insect population
Authors: McCoy Rajiv C. , Garud Nandita R. , Kelley Joanna L. , Boggs Carol L. , Petrov Dmitri A.
Published: November 2013

The analysis of molecular data from natural populations has allowed researchers to answer diverse ecological questions that were previously intractable. In particular, ecologists are often interested in the demographic history of populations, information that is rarely available from historical records. Methods have been developed to infer demographic parameters from genomic data, but it is not well understood how inferred parameters compare to true population history or depend on aspects of experimental design. Here, we present and evaluate a method ofSNPdiscovery usingRNAsequencing and demographic inference using the programδaδi, which uses a diffusion approximation to the allele frequency spectrum to fit demographic models. We test these methods in a population of the checkerspot butterflyEuphydryas gillettii. This population was intentionally introduced to Gothic, Colorado in 1977 and has as experienced extreme fluctuations including bottlenecks of fewer than 25 adults, as documented by nearly annual field surveys. UsingRNAsequencing of eight individuals from Colorado and eight individuals from a native population in Wyoming, we generate the first genomic resources for this system. While demographic inference is commonly used to examine ancient demography, our study demonstrates that our inexpensive, all‐in‐one approach to marker discovery and genotyping provides sufficient data to accurately infer the timing of a recent bottleneck. This demographic scenario is relevant for many species of conservation concern, few of which have sequenced genomes. Our results are remarkably insensitive to sample size or number of genomic markers, which has important implications for applying this method to other nonmodel systems.
Kin‐bias, breeding site selection and female fitness in a cannibalistic Neotropical frog
Authors: Muralidhar P. , de Sá F. P. , Haddad C. F. B. , Zamudio K. R.
Published: November 2013

Resource availability influences sexual selection within populations and determines whether behaviours such as territoriality or resource sharing are adaptive. In Thoropa taophora, a frog endemic to the Atlantic Coastal Rainforest of Brazil, males compete for and defend limited breeding sites while females often share breeding sites with other females; however, sharing breeding sites may involve costs due to cannibalism by conspecific tadpoles. We studied a breeding population of T. taophora to determine (i) whether this species exhibits polygynous mating involving female choice for territorial males and limited breeding resources; (ii) whether limited breeding resources create the potential for male–male cooperation in defence of neighbouring territories; and (iii) whether females sharing breeding sites exhibit kin‐biased breeding site choice, possibly driven by fitness losses due to cannibalism among offspring of females sharing sites. We used microsatellites to reconstruct parentage and quantify relatedness at eight breeding sites in our focal population, where these sites are scarce, and in a second population, where sites are abundant. We found that at localities where the appropriate sites for reproduction are spatially limited, the mating system for this species is polygynous, with typically two females sharing a breeding site with a male. We also found that females exhibit negative kin‐bias in their choice of breeding sites, potentially to maximize their inclusive fitness by avoiding tadpole cannibalism of highly related kin. Our results indicate that male territorial defence and female site sharing are likely important components of this mating system, and we propose that kinship‐dependent avoidance in mating strategies may be more general than previously realized.
Stretch‐Dependent Smooth Muscle Differentiation in the Portal Vein—Role of Actin Polymerization, Calcium Signaling, and microRNAs
Authors: Albinsson Sebastian , Bhattachariya Anirban , Hellstrand Per
Published: November 2013

The mechanical forces acting on SMC in the vascular wall are known to regulate processes such as vascular remodeling and contractile differentiation. However, investigations to elucidate the underlying mechanisms of mechanotransduction in smooth muscle have been hampered by technical limitations associated with mechanical studies on pressurized small arteries, due primarily to the small amount of available tissue. The murine portal vein is a relatively large vessel showing myogenic tone that in many respects recapitulates the properties of small resistance vessels. Studies on stretched portal veins to elucidate mechanisms of mechanotransduction in the vascular wall have shown that stretch‐sensitive regulation of contractile differentiation is mediated via Rho‐activation and actin polymerization, while stretch‐induced growth is regulated by the MAPK pathway. In this review, we have summarized findings on mechanotransduction in the portal vein with focus on stretch‐induced contractile differentiation and the role of calcium, actin polymerization and miRNAs in this response.
Transglutamination allows production and characterization of native‐sized ELPylated spider silk proteins from transgenic plants
Authors: Weichert Nicola , Hauptmann Valeska , Menzel Matthias , Schallau Kai , Gunkel Philip , Hertel Thomas C. , Pietzsch Markus , Spohn Uwe , Conrad Udo
Published: November 2013

In the last two decades it was shown that plants have a great potential for production of specific heterologous proteins. But high cost and inefficient downstream processing are a main technical bottleneck for the broader use of plant‐based production technology especially for protein‐based products, for technical use as fibres or biodegradable plastics and also for medical applications. High‐performance fibres from recombinant spider silks are, therefore, a prominent example. Spiders developed rather different silk materials that are based on proteins. These spider silks show excellent properties in terms of elasticity and toughness. Natural spider silk proteins have a very high molecular weight, and it is precisely this property which is thought to give them their strength. Transgenic plants were generated to produce ELPylated recombinant spider silk derivatives. These fusion proteins were purified byInverseTransitionCycling (ITC) and enzymatically multimerized with transglutaminasein vitro. Layers produced by casting monomers and multimers were characterized using atomic force microscopy (AFM) andAFM‐based nanoindentation. The layered multimers formed by mixing lysine‐ and glutamine‐tagged monomers were associated with the highest elastic penetration modulus.
BrpSPL9 (Brassica rapa ssp. pekinensis SPL9) controls the earliness of heading time in Chinese cabbage
Authors: Wang Yali , Wu Feijie , Bai Jinjuan , He Yuke
Published: November 2013

The leafy heads of cabbage (Brassica oleracea), Chinese cabbage (Brassica rapa ssp. pekinensis), Brussels sprouts (B. oleracea ssp. gemmifera) and lettuce (Lactuca sativa) comprise extremely incurved leaves that are edible vegetable products. The heading time is important for high quality and yield of these crops. Here, we report that BrpSPL9‐2 (B. rapa ssp. pekinensis SQUAMOSA PROMOTER BINDING‐LIKE 9‐2), a target gene of microRNA brp‐miR156, controls the heading time of Chinese cabbage. Quantitative measurements of leaf shapes, sizes, colour and curvature indicated that heading is a late adult phase of vegetative growth. During the vegetative period, miR156 levels gradually decreased from the seedling stage to the heading one, whereas BrpSPL9‐2 and BrpSPL15‐1 mRNAs increased progressively and reached the highest levels at the heading stage. Overexpression of a mutated miR156‐resistant form of BrpSPL9‐2 caused the significant earliness of heading, concurrent with shortening of the seedling and rosette stages. By contrast, overexpression of miR156 delayed the folding time, concomitant with prolongation of the seedling and rosette stages. Morphological analysis reveals that the significant earliness of heading in the transgenic plants overexpressing BrpSPL9‐2 gene was produced because the juvenile phase was absent and the early adult phase shortened, whereas the significant delay of folding in the transgenic plants overexpressing Brp‐MIR156a was due to prolongation of the juvenile and early adult phases. Thus, miR156 and BrpSPL9 genes are potentially important for genetic improvement of earliness of Chinese cabbage and other crops.
Inter‐instrument comparison of particle‐size analysers
Authors: Roberson Sam , Weltje Gert Jan
Published: November 2013

This paper presents a methodological framework for inter‐instrument comparison of different particle‐size analysers. The framework consists of: (i) quantifying the difference between complete particle‐size distributions; (ii) identifying the best regression model for homogenizing data sets of particle‐size distributions measured by different instruments; (iii) quantifying the precision of a range of particle‐size analysers; and (iv) identifying the most appropriate instrument for analysing a given set of samples. Thelog‐ratiotransform is applied to particle‐size distributions throughout this study to avoid the pitfalls of analysing percentage‐frequency data in ‘closed‐space’. ANormalized Distancestatistic is used to quantify the difference between particle‐size distributions and assess the performance oflog‐ratioregression models. Forty‐six different regression models are applied to sediment samples measured by both sieve‐pipette and laser analysis. Interactive quadratic regression models offer the best means of homogenizing data sets of particle‐size distributions measured by different instruments into a comparable format. However, quadratic interactive log‐ratio regression models require a large number of training samples (n > 80) to achieve optimal performance compared to linear regression models (n = 50). The precision of ten particle‐size analysis instruments was assessed using a data set of ten replicate measurements made of four previously published silty sediment samples. Instrument precision is quantified as the medianNormalized Differencemeasured between the ten replicate measurements made for each sediment sample. TheDifferentiation Powerstatistic is introduced to assess the ability of each instrument to detect differences between the four sediment samples.Differentiation Powerscores show that instruments based on laser diffraction principles are able to differentiate most effectively between the samples of silty sediment at a 95% confidence level. Instruments applying the principles of sedimentation offer the next most precise approach.
Keys for microRNA expression profiling of single rat hypothalamic nuclei and multiplex sequencing strategies
Authors: Baroin‐Tourancheau A. , Benigni X. , Benoit C. , Doubi‐Kadmiri S. , Vacher C. M. , Taouis M. , Amar L.
Published: November 2013

New Findings

What is the topic of this review?  We sought to establish a high number of microRNA expression profiles of individual hypothalamic nuclei of single rats.

What advances does it highlight?  This can be achieved at a cost compatible with laboratory finance by using tagged cDNA libraries constructed from purified small RNAs and a multiplex sequencing strategy.

Integrative research has taken on the challenge of addressing questions in physiology by using novel knowledge and novel techniques. Recently, small and long non‐coding RNAs have emerged as key regulators of gene expression, while next‐generation sequencing technologies have revolutionized the characterization of genomes and gene expression. For a decade, it has been known that microRNAs (miRNAs) are RNAs of 18–24 bases that regulate gene expression in mammals. Here, we first describe the nature of miRNAs and the advantages of high‐throughput sequencing technologies for establishing miRNA expression profiles. The hypothalamus harbours a dozen specialized areas or nuclei, the sampling of which is required to establish physiologically relevant miRNA expression profiles. MicroRNA expression profiling from single animals is also important for investigating potential genetic or epigenetic differences between individuals. Establishing a large number of miRNA expression profiles of individual hypothalamic nuclei of single rats at a cost compatible with laboratory finance can be achieved by using tagged cDNA libraries constructed from purified small RNAs and a multiplex sequencing strategy. We continue this report by surveying specificities of the different strategies that are used at present for constructing tagged cDNA libraries and provide a comparative analysis of miRNA expression profiles from hypothalamic arcuate nuclei of seven male Wistar rats.
AltitudeOmics: effect of ascent and acclimatization to 5260 m on regional cerebral oxygen delivery
Authors: Subudhi Andrew W. , Fan Jui‐Lin , Evero Oghenero , Bourdillon Nicolas , Kayser Bengt , Julian Colleen G. , Lovering Andrew T. , Roach Robert C.
Published: November 2013

New Findings

What is the central question of this study?

Hypoxia associated with ascent to high altitude may threaten cerebral oxygen delivery. We sought to determine whether there are regional changes in the distribution of cerebral blood flow that might favour oxygen delivery to areas associated with basic homeostatic functions to promote survival in this extreme environment.

What is the main finding and its importance?

We show evidence of a ‘brain‐sparing’ effect during acute exposure to high altitude, in which there is a slight increase in relative oxygen delivery to the posterior cerebral circulation. This may serve to support basic regulatory functions associated with the brainstem and hypothalamus.

Cerebral hypoxaemia associated with rapid ascent to high altitude can be life threatening; yet, with proper acclimatization, cerebral function can be maintained well enough for humans to thrive. We investigated adjustments in global and regional cerebral oxygen delivery () as 21 healthy volunteers rapidly ascended and acclimatized to 5260 m. Ultrasound indices of cerebral blood flow in internal carotid and vertebral arteries were measured at sea level, upon arrival at 5260 m (ALT1; atmospheric pressure 409 mmHg) and after 16 days of acclimatization (ALT16). Cerebral was calculated as the product of arterial oxygen content and flow in each respective artery and summed to estimate global cerebral blood flow. Vascular resistances were calculated as the quotient of mean arterial pressure and respective flows. Global cerebral blood flow increased by ∼70% upon arrival at ALT1 (P < 0.001) and returned to sea‐level values at ALT16 as a result of changes in cerebral vascular resistance. A reciprocal pattern in arterial oxygen content maintained global cerebral throughout acclimatization, although to the posterior cerebral circulation was increased by ∼25% at ALT1 (P = 0.032). We conclude that cerebral is well maintained upon acute exposure and acclimatization to hypoxia, particularly in the posterior and inferior regions of the brain associated with vital homeostatic functions. This tight regulation of cerebral was achieved through integrated adjustments in local vascular resistances to alter cerebral perfusion during both acute and chronic exposure to hypoxia.
Dissociation between blood pressure and heart rate response to hypoxia after bilateral carotid body removal in men with systolic heart failure
Authors: Niewinski Piotr , Janczak Dariusz , Rucinski Artur , Tubek Stanislaw , Engelman Zoar J. , Jazwiec Przemyslaw , Banasiak Waldemar , Sobotka Paul A. , Hart Emma C. J. , Paton Julian F. R. , Ponikowski Piotr
Published: November 2013

New Findings

What is the central question of this study?

Carotid body denervation removes the ventilatory response to acute hypoxia, although haemodynamic responses to acute hypoxia after carotid body removal have not been described conclusively in humans.

What is the main finding and its importance?

Carotid body removal results in dissociation of heart rate and blood pressure responses to hypoxia in human subjects. While the heart rate response (tachycardia) is maintained, there is a significant attenuation of the blood pressure response (hypertension), which indicates the existence of different sensory afferent pathways in the haemodynamic response to hypoxia that has important clinical implications for this novel therapeutic modality.

While the ventilatory response to hypoxia is known to be mediated by the carotid bodies, the origin of the haemodynamic alterations evoked by hypoxia is less certain. Bilateral carotid body removal (CBR) performed to treat congestive heart failure may serve as a model to improve our understanding of haemodynamic responses to hypoxia in humans. We studied six congestive heart failure patients before and 1 month after CBR [median (interquartile range): age, 58.5 (56–61) years old; and ejection fraction, 32 (25–34)%]. Peripheral chemosensitivity (hypoxic ventilatory response) was equated to the slope relating lowest oxygen saturation to highest minute ventilation following exposures to hypoxia. Likewise, systolic blood pressure (SBP), diastolic blood pressure (DBP) and heart rate (HR) slopes were calculated as slopes relating the lowest oxygen saturations to the highest SBP, DBP and HR responses. We found that CBR reduces the hypoxic ventilatory response (∆91%, P < 0.05), SBP (∆71%, P < 0.05) and DBP slopes (∆59%, P = 0.07). In contrast, the HR slope remained unchanged. The dissociation between the blood pressure and HR responses after CBR shows involvement of a different chemoreceptive site(s) maintaining the response to acute hypoxia. We conclude that carotid bodies are responsible for ventilatory and blood pressure responses, while the HR response might be mediated by the aortic bodies. The significant reduction of the blood pressure response to hypoxia after CBR suggests a decrease in sympathetic tone, which is of particular clinical relevance in congestive heart failure.
Differential visceral pain sensitivity and colonic morphology in four common laboratory rat strains
Authors: O'Malley Dervla , Julio‐Pieper Marcela , O'Mahony Siobhain M. , Dinan Timothy G. , Cryan John F.
Published: November 2013

New Findings

What is the central question of this study?

Does stress sensitivity and susceptibility to inflammation innate to certain rat strains make them vulnerable to bowel dysfunction?

What is the main finding and its importance?

Of four different rat strains, the Lewis rat, which displays both susceptibility to gastrointestinal inflammation and sensitivity to stress, exhibits the most aberrant gastrointestinal morphology and visceral pain sensitivity. Given the similarities to human functional bowel disorders, such as irritable bowel syndrome, this may make it a good model of this disease.

Irritable bowel syndrome is a common, debilitating gastrointestinal (GI) disorder characterized by episodic exacerbations of symptoms such as abdominal pain, bloating and altered bowel habit. Contributory factors for the development of irritable bowel syndrome include genetics, childhood trauma and prior GI infection leading to chronic low‐grade inflammation or immune activation. Additional considerations in comprehending the chronic relapsing pattern that typifies irritable bowel syndrome symptoms are the effects of both psychosocial and infection‐related stresses. Background stress and immune profiles can influence gut permeability and visceral pain sensitivity. This study examined whether innate susceptibility to inflammation and stress sensitivity in four rat strains is associated with bowel dysfunction. The pain threshold to colorectal distension was assessed in Lewis, Fischer (F344) and spontaneously hypertensive rats and compared with Sprague–Dawley control animals. Colons were subsequently excised and morphologically assessed for total length, goblet cell hyperplasia and muscle and mucosal thickness. Lewis rats displayed visceral hypersensitivity compared with other strains. At a morphological level, the gastrointestinal tract from these rats displayed mucosal goblet cell hyperplasia and alterations in muscle layer thickness. The Lewis rat strain, which is reported to have increased susceptibility to GI inflammation in addition to stress sensitivity, had the most prominent features of physiological and morphological GI dysfunction. These data support the hypothesis that background strain is a key factor in the development and exacerbation of bowel dysfunction in rodent models.
Competition for Ammonia Influences the Structure of Chemotrophic Communities in Geothermal Springs
Authors: Hamilton Trinity L. , Koonce Evangeline , Howells Alta , Havig Jeff R. , Jewell Talia , de la Torre José R. , Peters John W. , Boyd Eric S.
Published: November 2013

Source waters sampled from Perpetual Spouter hot spring (pH 7.03, 86.4°C), Yellowstone National Park, WY, have low concentrations of total ammonia, nitrite, and nitrate, suggesting nitrogen (N) limitation and/or tight coupling of N cycling processes. Dominant small-subunit rRNA sequences in Perpetual Spouter source sediments are closely affiliated with the ammonia-oxidizing archaeon “
Candidatus
Nitrosocaldus yellowstonii” and the putatively nitrogen-fixing (diazotrophic) bacterium
Thermocrinis albus
, respectively, suggesting that these populations may interact at the level of the bioavailable N pool, specifically, ammonia. This hypothesis was evaluated by using a combination of geochemical, physiological, and transcriptomic analyses of sediment microcosms. Amendment of microcosms with allylthiourea, an inhibitor of ammonia oxidation, decreased rates of acetylene reduction (a proxy for N
2
fixation) and nitrite production (a proxy for ammonia oxidation) and decreased transcript levels of structural genes involved in both nitrogen fixation (
nifH
) and ammonia oxidation (
amoA
). In contrast, amendment of microcosms with ammonia stimulated nitrite production and increased
amoA
transcript levels while it suppressed rates of acetylene reduction and decreased
nifH
transcript levels. Sequencing of amplified
nifH
and
amoA
transcripts from native sediments, as well as microcosms, at 2 and 4 h postamendment, indicates that the dominant and responsive populations involved in ammonia oxidation and N
2
fixation are closely affiliated with
Ca
. Nitrosocaldus yellowstonii and
T. albus
, respectively. Collectively, these results suggest that ammonia-oxidizing archaea, such as
Ca
. Nitrosocaldus yellowstonii, have an apparent affinity for ammonia that is higher than that of the diazotrophs present in this ecosystem. Depletion of the bioavailable N pool through the activity of ammonia-oxidizing archaea likely represents a strong selective pressure for the inclusion of organisms capable of nitrogen fixation in geothermal communities. These observations help to explain the strong pattern in the codistribution of ammonia-oxidizing archaea and diazotrophs in circumneutral-to-alkaline geothermal springs.
Interspecies Interactions Determine the Impact of the Gut Microbiota on Nutrient Allocation in Drosophila melanogaster
Authors: Newell Peter D. , Douglas Angela E.
Published: November 2013

The animal gut is perpetually exposed to microorganisms, and this microbiota affects development, nutrient allocation, and immune homeostasis. A major challenge is to understand the contribution of individual microbial species and interactions among species in shaping these microbe-dependent traits. Using the
Drosophila melanogaster
gut microbiota, we tested whether microbe-dependent performance and nutritional traits of
Drosophila
are functionally modular, i.e., whether the impact of each microbial taxon on host traits is independent of the presence of other microbial taxa. Gnotobiotic flies were constructed with one or a set of five of the
Acetobacter
and
Lactobacillus
species which dominate the gut microbiota of conventional flies (
Drosophila
with untreated microbiota). Axenic (microbiota-free) flies exhibited prolonged development time and elevated glucose and triglyceride contents. The low glucose content of conventional flies was recapitulated in gnotobiotic
Drosophila
flies colonized with any of the 5 bacterial taxa tested. In contrast, the development rates and triglyceride levels in monocolonized flies varied depending on the taxon present:
Acetobacter
species supported the largest reductions, while most
Lactobacillus
species had no effect. Only flies with both
Acetobacter
and
Lactobacillus
had triglyceride contents restored to the level in conventional flies. This could be attributed to two processes:
Lactobacillus
-mediated promotion of
Acetobacter
abundance in the fly and a significant negative correlation between fly triglyceride content and
Acetobacter
abundance. We conclude that the microbial basis of host traits varies in both specificity and modularity; microbe-mediated reduction in glucose is relatively nonspecific and modular, while triglyceride content is influenced by interactions among microbes.
pH-Dependent Activation of Streptomyces hygroscopicus Transglutaminase Mediated by Intein
Authors: Du Kun , Liu Zhongmei , Cui Wenjing , Zhou Li , Liu Yi , Du Guocheng , Chen Jian , Zhou Zhemin
Published: November 2013

Microbial transglutaminase (MTG) from
Streptomyces
is naturally secreted as a zymogen (pro-MTG), which is then activated by the removal of its N-terminal proregion by additional proteases. Inteins are protein-intervening sequences that catalyze protein splicing without cofactors. In this study, a pH-dependent
Synechocystis
sp. strain PCC6803 DnaB mini-intein (SDB) was introduced into pro-MTG to simplify its activation process by controlling pH. The recombinant protein (pro-SDB-MTG) was obtained, and the activation process was determined to take 24 h at pH 7
in vitro
. To investigate the effect of the first residue in MTG on the activity and the cleavage time, two variants, pro-SDB-MTG(D1S) and pro-SDB-MTG(ΔD1), were expressed, and the activation time was found to be 6 h and 30 h, respectively. The enzymatic property and secondary structure of the recombinant MTG and two variants were similar to those of the wild type, indicating that the insertion of mini-intein did not affect the function of MTG. This insignificant effect was further illustrated by molecular dynamics simulations. This study revealed a controllable and effective strategy to regulate the activation process of pro-MTG mediated by a mini-intein, and it may have great potential for industrial MTG production.
Oral Bacterial Communities in Individuals with Type 2 Diabetes Who Live in Southern Thailand
Authors: Kampoo Kanokporn , Teanpaisan Rawee , Ledder Ruth G. , McBain Andrew J.
Published: November 2013

Type 2 diabetes mellitus is increasingly common in Thailand and elsewhere. In the present investigation, the bacteriological composition of saliva and supragingival plaque in Thai diabetics with and without active dental caries and in nondiabetics was determined by differential culture and eubacterial DNA profiling. Potential associations between fasting blood sugar and glycosylated hemoglobin (biomarkers of current and historical glucose control, respectively) with decayed, missing, and filled teeth and with salivary
Streptococcus
and
Lactobacillus
counts were also investigated. The incidence of active dental caries was greater in the Thai diabetics than in nondiabetics, and the numbers of total streptococci and lactobacilli were significantly higher in supragingival plaque from diabetics than in nondiabetics.
Lactobacillus
counts in the saliva and supragingival plaque of diabetics with active caries were significantly higher than those in diabetics without active caries. Oral eubacterial DNA profiles of diabetic versus nondiabetic individuals and of diabetics with active caries versus those without active caries could not be readily differentiated through cluster analysis or multidimensional scaling. The elevated caries incidence in the Thai diabetics was positively associated with numbers of bacteria of the acidogenic/acid-tolerant genera
Streptococcus
and
Lactobacillus. Lactobacillus
bacterial numbers were further elevated in diabetics with active caries, although salivary eubacterial DNA profiles were not significantly altered.
Expression and Characterization of
CYP52
Genes Involved in the Biosynthesis of Sophorolipid and Alkane Metabolism from Starmerella bombicola
Authors: Huang Fong-Chin , Peter Alyssa , Schwab Wilfried
Published: November 2013

Three cytochrome P450 monooxygenase
CYP52
gene family members were isolated from the sophorolipid-producing yeast
Starmerella bombicola
(former
Candida bombicola
), namely,
CYP52E3
,
CYP52M1
, and
CYP52N1
, and their open reading frames were cloned into the pYES2 vector for expression in
Saccharomyces cerevisiae
. The functions of the recombinant proteins were analyzed with a variety of alkane and fatty acid substrates using microsome proteins or a whole-cell system. CYP52M1 was found to oxidize C
16
to C
20
fatty acids preferentially. It converted oleic acid (C
18:1
) more efficiently than stearic acid (C
18:0
) and linoleic acid (C
18:2
) and much more effectively than α-linolenic acid (C
18:3
). No products were detected when C
10
to C
12
fatty acids were used as the substrates. Moreover, CYP52M1 hydroxylated fatty acids at their ω- and ω-1 positions. CYP52N1 oxidized C
14
to C
20
saturated and unsaturated fatty acids and preferentially oxidized palmitic acid, oleic acid, and linoleic acid. It only catalyzed ω-hydroxylation of fatty acids. Minor ω-hydroxylation activity against myristic acid, palmitic acid, palmitoleic acid, and oleic acid was shown for CYP52E3. Furthermore, the three P450s were coassayed with glucosyltransferase UGTA1. UGTA1 glycosylated all hydroxyl fatty acids generated by CYP52E3, CYP52M1, and CYP52N1. The transformation efficiency of fatty acids into glucolipids by CYP52M1/UGTA1 was much higher than those by CYP52N1/UGTA1 and CYP52E3/UGTA1. Taken together, CYP52M1 is demonstrated to be involved in the biosynthesis of sophorolipid, whereas CYP52E3 and CYP52N1 might be involved in alkane metabolism in
S. bombicola
but downstream of the initial oxidation steps.
From Genus to Phylum: Large-Subunit and Internal Transcribed Spacer rRNA Operon Regions Show Similar Classification Accuracies Influenced by Database Composition
Authors: Porras-Alfaro Andrea , Liu Kuan-Liang , Kuske Cheryl R. , Xie Gary
Published: November 2013

We compared the classification accuracy of two sections of the fungal internal transcribed spacer (ITS) region, individually and combined, and the 5′ section (about 600 bp) of the large-subunit rRNA (LSU), using a naive Bayesian classifier and BLASTN. A hand-curated ITS-LSU training set of 1,091 sequences and a larger training set of 8,967 ITS region sequences were used. Of the factors evaluated, database composition and quality had the largest effect on classification accuracy, followed by fragment size and use of a bootstrap cutoff to improve classification confidence. The naive Bayesian classifier and BLASTN gave similar results at higher taxonomic levels, but the classifier was faster and more accurate at the genus level when a bootstrap cutoff was used. All of the ITS and LSU sections performed well (>97.7% accuracy) at higher taxonomic ranks from kingdom to family, and differences between them were small at the genus level (within 0.66 to 1.23%). When full-length sequence sections were used, the LSU outperformed the ITS1 and ITS2 fragments at the genus level, but the ITS1 and ITS2 showed higher accuracy when smaller fragment sizes of the same length and a 50% bootstrap cutoff were used. In a comparison using the larger ITS training set, ITS1 and ITS2 had very similar accuracy classification for fragments between 100 and 200 bp. Collectively, the results show that any of the ITS or LSU sections we tested provided comparable classification accuracy to the genus level and underscore the need for larger and more diverse classification training sets.
A Moderately Thermophilic Mixed Microbial Culture for Bioleaching of Chalcopyrite Concentrate at High Pulp Density
Authors: Wang Yuguang , Zeng Weimin , Qiu Guanzhou , Chen Xinhua , Zhou Hongbo
Published: November 2013

Three kinds of samples (acid mine drainage, coal mine wastewater, and thermal spring) derived from different sites were collected in China. Thereafter, these samples were combined and then inoculated into a basal salts solution in which different substrates (ferrous sulfate, elemental sulfur, and chalcopyrite) served as energy sources. After that, the mixed cultures growing on different substrates were pooled equally, resulting in a final mixed culture. After being adapted to gradually increasing pulp densities of chalcopyrite concentrate by serial subculturing for more than 2 years, the final culture was able to efficiently leach the chalcopyrite at a pulp density of 20% (wt/vol). At that pulp density, the culture extracted 60.4% of copper from the chalcopyrite in 25 days. The bacterial and archaeal diversities during adaptation were analyzed by denaturing gradient gel electrophoresis and constructing clone libraries of the 16S rRNA gene. The results show that the culture consisted mainly of four species, including
Leptospirillum ferriphilum
,
Acidithiobacillus caldus
,
Sulfobacillus acidophilus
, and
Ferroplasma thermophilum
, before adapting to a pulp density of 4%. However,
L. ferriphilum
could not be detected when the pulp density was greater than 4%. Real-time quantitative PCR was employed to monitor the microbial dynamics during bioleaching at a pulp density of 20%. The results show that
A. caldus
was the predominant species in the initial stage, while
S. acidophilus
rather than
A. caldus
became the predominant species in the middle stage.
F. thermophilum
accounted for the greatest proportion in the final stage.
Identification and Environmental Distribution ofdcpA, Which Encodes the Reductive Dehalogenase Catalyzing the Dichloroelimination of 1,2-Dichloropropane to Propene in Organohalide-Respiring Chloroflexi
Authors: Padilla-Crespo Elizabeth , Yan Jun , Swift Cynthia , Wagner Darlene D. , Chourey Karuna , Hettich Robert L. , Ritalahti Kirsti M. , Löffler Frank E.
Published: November 2013

Dehalococcoides mccartyistrains KS and RC grow with 1,2-dichloropropane (1,2-D) as an electron acceptor in enrichment cultures derived from hydrocarbon-contaminated and pristine river sediments, respectively. Transcription, expression, enzymatic, and PCR analyses implicated the reductive dehalogenase genedcpAin 1,2-D dichloroelimination to propene and inorganic chloride. Quantitative real-time PCR (qPCR) analyses demonstrated aD. mccartyicell increase during growth with 1,2-D and suggested that bothD. mccartyistrains carried a singledcpAgene copy per genome.D. mccartyistrain RC and strain KS produced 1.8 × 107± 0.1 × 107and 1.4 × 107± 0.5 × 107cells per μmol of propene formed, respectively. ThedcpAgene was identified in 1,2-D-to-propene-dechlorinating microcosms established with sediment samples collected from different geographical locations in Europe and North and South America. Clone library analysis revealed two distinctdcpAphylogenetic clusters, both of which were captured by thedcpAgene-targeted qPCR assay, suggesting that the qPCR assay is useful for site assessment and bioremediation monitoring at 1,2-D-contaminated sites.
New Model for Electron Flow for Sulfate Reduction in Desulfovibrio alaskensis G20
Authors: Keller Kimberly L. , Rapp-Giles Barbara J. , Semkiw Elizabeth S. , Porat Iris , Brown Steven D. , Wall Judy D.
Published: November 2013

To understand the energy conversion activities of the anaerobic sulfate-reducing bacteria, it is necessary to identify the components involved in electron flow. The importance of the abundant type I tetraheme cytochrome
c
3
(TpI
c
3
) as an electron carrier during sulfate respiration was questioned by the previous isolation of a null mutation in the gene encoding TpI
c
3
,
cycA
, in
Desulfovibrio alaskensis
G20. Whereas respiratory growth of the CycA mutant with lactate and sulfate was little affected, growth with pyruvate and sulfate was significantly impaired. We have explored the phenotype of the CycA mutant through physiological tests and transcriptomic and proteomic analyses. Data reported here show that electrons from pyruvate oxidation do not reach adenylyl sulfate reductase, the enzyme catalyzing the first redox reaction during sulfate reduction, in the absence of either CycA or the type I cytochrome
c
3
:menaquinone oxidoreductase transmembrane complex, QrcABCD. In contrast to the wild type, the CycA and QrcA mutants did not grow with H
2
or formate and sulfate as the electron acceptor. Transcriptomic and proteomic analyses of the CycA mutant showed that transcripts and enzymes for the pathway from pyruvate to succinate were strongly decreased in the CycA mutant regardless of the growth mode. Neither the CycA nor the QrcA mutant grew on fumarate alone, consistent with the omics results and a redox regulation of gene expression. We conclude that TpI
c
3
and the Qrc complex are
D. alaskensis
components essential for the transfer of electrons released in the periplasm to reach the cytoplasmic adenylyl sulfate reductase and present a model that may explain the CycA phenotype through confurcation of electrons.
UV Light Inactivation of Human and Plant Pathogens in Unfiltered Surface Irrigation Water
Authors: Jones Lisa A. , Worobo Randy W. , Smart Christine D.
Published: November 2013

Fruit and vegetable growers continually battle plant diseases and food safety concerns. Surface water is commonly used in the production of fruits and vegetables and can harbor both human- and plant-pathogenic microorganisms that can contaminate crops when used for irrigation or other agricultural purposes. Treatment methods for surface water are currently limited, and there is a need for suitable treatment options. A liquid-processing unit that uses UV light for the decontamination of turbid juices was analyzed for its efficacy in the treatment of surface waters contaminated with bacterial or oomycete pathogens, i.e.,
Escherichia coli
,
Salmonella enterica
,
Listeria monocytogenes
,
Clavibacter michiganensis
subsp.
michiganensis
,
Pseudomonas syringae
pv. tomato, and
Phytophthora capsici
. Five-strain cocktails of each pathogen, containing approximately 10
8
or 10
9
CFU/liter for bacteria or 10
4
or 10
5
zoospores/liter for
Ph. capsici
, were inoculated into aliquots of two turbid surface water irrigation sources and processed with the UV unit. Pathogens were enumerated before and after treatment. In general, as the turbidity of the water source increased, the effectiveness of the UV treatment decreased, but in all cases, 99.9% or higher inactivation was achieved. Log reductions ranged from 10.0 to 6.1 and from 5.0 to 4.2 for bacterial pathogens and
Ph. capsici
, respectively.
Interaction of Aeromonas Strains with Lactic Acid Bacteria via Caco-2 Cells
Authors: Hatje E. , Neuman C. , Katouli M.
Published: November 2013

The genus
Aeromonas
includes some species that have now been identified as human pathogens of significant medical importance. We investigated the ability of 13 selected
Aeromonas
strains belonging to nine species isolated from clinical cases (
n
= 5), environmental waters (
n
= 5), and fish (
n
= 3) to adhere to and translocate Caco-2 cells in the absence and presence of two lactic acid bacteria (LAB), i.e.,
Lactobacillus acidophilus
and
Bifidobacterium breve
.
Aeromonas
isolates were also assessed for their cytotoxicity, the presence of virulence genes, and hemolysin production. Among the clinical isolates, one strain of
Aeromonas veronii
biovar veronii and two strains of
Aeromonas hydrophila
carried cytotoxin (
act
), heat-labile toxin (
alt
), hemolysin (
hlyA
), and aerolysin (
aerA
) genes, were cytotoxic to Vero cells, produced hemolysin, and showed higher adherence to Caco-2 cells. In contrast, this was seen in only one environmental strain, a strain of
A. veronii
biovar sobria. When
Aeromonas
strains were coinoculated with LAB onto Caco-2 cells, their level of adhesion was reduced. However, their rate of translocation in the presence of LAB increased and was significantly (
P
< 0.05) higher among fish strains. We suggest that either the interaction between
Aeromonas
and LAB strains could have a detrimental effect on the Caco-2 cells, allowing the
Aeromonas
to translocate more readily, or the presence of the LAB stimulated the
Aeromonas
strains to produce more toxins and/or increase their translocation rate.
Bifidobacterium bifidum PRL2010 Modulates the Host Innate Immune Response
Authors: Turroni Francesca , Taverniti Valentina , Ruas-Madiedo Patricia , Duranti Sabrina , Guglielmetti Simone , Lugli Gabriele Andrea , Gioiosa Laura , Palanza Paola , Margolles Abelardo , van Sinderen Douwe , Ventura Marco
Published: November 2013

Here, we describe data obtained from transcriptome profiling of human cell lines and intestinal cells of a murine model upon exposure and colonization, respectively, withBifidobacterium bifidumPRL2010. Significant changes were detected in the transcription of genes that are known to be involved in innate immunity. Furthermore, results from enzyme-linked immunosorbent assays (ELISAs) showed that exposure toB. bifidumPRL2010 causes enhanced production of interleukin 6 (IL-6) and IL-8 cytokines, presumably through NF-κB activation. The obtained global transcription profiles strongly suggest thatBifidobacterium bifidumPRL2010 modulates the innate immune response of the host.
The Fitness Advantage of Commercial Wine Yeasts in Relation to the Nitrogen Concentration, Temperature, and Ethanol Content under Microvinification Conditions
Authors: García-Ríos Estéfani , Gutiérrez Alicia , Salvadó Zoel , Arroyo-López Francisco Noé , Guillamon José Manuel
Published: November 2013

The effect of the main environmental factors governing wine fermentation on the fitness of industrial yeast strains has barely received attention. In this study, we used the concept of fitness advantage to measure how increasing nitrogen concentrations (0 to 200 mg N/liter), ethanol (0 to 20%), and temperature (4 to 45°C) affects competition among four commercial wine yeast strains (PDM, ARM, RVA, and TTA). We used a mathematical approach to model the hypothetical time needed for the control strain (PDM) to out-compete the other three strains in a theoretical mixed population. The theoretical values obtained were subsequently verified by competitive mixed fermentations in both synthetic and natural musts, which showed a good fit between the theoretical and experimental data. Specifically, the data show that the increase in nitrogen concentration and temperature values improved the fitness advantage of the PDM strain, whereas the presence of ethanol significantly reduced its competitiveness. However, the RVA strain proved to be the most competitive yeast for the three enological parameters assayed. The study of the fitness of these industrial strains is of paramount interest for the wine industry, which uses them as starters of their fermentations. Here, we propose a very simple method to model the fitness advantage, which allows the prediction of the competitiveness of one strain with respect to different abiotic factors.