Host Plant Associations of an Entomopathogenic Variety of the Fungus, Colletotrichum acutatum, Recovered from the Elongate Hemlock Scale, Fiorinia externa

A fungal epizootic has been detected in populations of the scale Fiorinia externa Ferris (Hemiptera: Diaspididae) in the eastern hemlock, Tsuga canadensis (L.) Carrière (Pinales: Pinaceae), of several northeastern states. Colletotrichum acutatum Simmonds var. fioriniae Marcelino and Gouli var. nov. inedit (Phyllachorales: Phyllachoraceae), a well-known plant pathogen, was the most commonly recovered fungus from these infected scales. This is the second report of a Colletotrichum sp. infecting scale insects. In Brazil C. gloeosporioides f. sp. ortheziidae recovered from Orthezia praelonga is under development as a biopesticide for citrus production. C. acutatum was detected growing endophytically in 28 species of plants within the epizootic areas. DNA sequences of the High Mobility Box at the MAT 1–2, mating type gene indicate that Colletotrichum sp. isolates recovered from scale insects and plants within epizootic areas were identical. Results from plant bioassays showed that this entomopathogenic Colletotrichum variety grew endophytically in all of the plants tested without causing external symptoms or signs of infection, with the exception of strawberry plants where mild symptoms of infection were observed. The implications of these findings with respect to the use of this fungus as a biological control agent are discussed

Characterizing low affinity epibatidine binding to α4β2 nicotinic acetylcholine receptors with ligand depletion and nonspecific binding

<p>Abstract</p> <p>Background</p> <p>Along with high affinity binding of epibatidine (<it>K</it>_d1≈10 pM) to α4β2 nicotinic acetylcholine receptor (nAChR), low affinity binding of epibatidine (<it>K</it>_d2≈1-10 nM) to an independent binding site has been reported. Studying this low affinity binding is important because it might contribute understanding about the structure and synthesis of α4β2 nAChR. The binding behavior of epibatidine and α4β2 AChR raises a question about interpreting binding data from two independent sites with ligand depletion and nonspecific binding, both of which can affect equilibrium binding of [³H]epibatidine and α4β2 nAChR. If modeled incorrectly, ligand depletion and nonspecific binding lead to inaccurate estimates of binding constants. Fitting total equilibrium binding as a function of total ligand accurately characterizes a single site with ligand depletion and nonspecific binding. The goal of this study was to determine whether this approach is sufficient with two independent high and low affinity sites.</p> <p>Results</p> <p>Computer simulations of binding revealed complexities beyond fitting total binding for characterizing the second, low affinity site of α4β2 nAChR. First, distinguishing low-affinity specific binding from nonspecific binding was a potential problem with saturation data. Varying the maximum concentration of [³H]epibatidine, simultaneously fitting independently measured nonspecific binding, and varying α4β2 nAChR concentration were effective remedies. Second, ligand depletion helped identify the low affinity site when nonspecific binding was significant in saturation or competition data, contrary to a common belief that ligand depletion always is detrimental. Third, measuring nonspecific binding without α4β2 nAChR distinguished better between nonspecific binding and low-affinity specific binding under some circumstances of competitive binding than did presuming nonspecific binding to be residual [³H]epibatidine binding after adding a large concentration of cold competitor. Fourth, nonspecific binding of a heterologous competitor changed estimates of high and low inhibition constants but did not change the ratio of those estimates.</p> <p>Conclusions</p> <p>Investigating the low affinity site of α4β2 nAChR with equilibrium binding when ligand depletion and nonspecific binding are present likely needs special attention to experimental design and data interpretation beyond fitting total binding data. Manipulation of maximum ligand and receptor concentrations and intentionally increasing ligand depletion are potentially helpful approaches.</p

Conifers in cold environments synchronize maximum growth rate of tree-ring formation with day length

Intra-annual radial growth rates and durations in trees are reported to differ greatly in relation to species, site and environmental conditions. However, very similar dynamics of cambial activity and wood formation are observed in temperate and boreal zones. Here, we compared weekly xylem cell production and variation in stem circumference in the main northern hemisphere conifer species (genera Picea, Pinus, Abies and Larix) from 1996 to 2003. Dynamics of radial growth were modeled with a Gompertz function, defining the upper asymptote (A), x-axis placement (β) and rate of change (κ). A strong linear relationship was found between the constants β and κ for both types of analysis. The slope of the linear regression, which corresponds to the time at which maximum growth rate occurred, appeared to converge towards the summer solstice. The maximum growth rate occurred around the time of maximum day length, and not during the warmest period of the year as previously suggested. The achievements of photoperiod could act as a growth constraint or a limit after which the rate of tree-ring formation tends to decrease, thus allowing plants to safely complete secondary cell wall lignification before winter

Examining the effects of sodium ions on the binding of antagonists to dopamine D2 and D3 receptors

Many G protein-coupled receptors have been shown to be sensitive to the presence of sodium ions (Na+). Using radioligand competition binding assays, we have examined and compared the effects of sodium ions on the binding affinities of a number of structurally diverse ligands at human dopamine D2 and dopamine D3 receptor subtypes, which are important therapeutic targets for the treatment of psychotic disorders. At both receptors, the binding affinities of the antagonists/inverse agonists SB-277011-A, L,741,626, GR 103691 and U 99194 were higher in the presence of sodium ions compared to those measured in the presence of the organic cation, N-methyl-D-glucamine, used to control for ionic strength. Conversely, the affinities of spiperone and (+)-butaclamol were unaffected by the presence of sodium ions. Interestingly, the binding of the antagonist/inverse agonist clozapine was affected by changes in ionic strength of the buffer used rather than the presence of specific cations. Similar sensitivities to sodium ions were seen at both receptors, suggesting parallel effects of sodium ion interactions on receptor conformation. However, no clear correlation between ligand characteristics, such as subtype selectivity, and sodium ion sensitivity were observed. Therefore, the properties which determine this sensitivity remain unclear. However these findings do highlight the importance of careful consideration of assay buffer composition for in vitro assays and when comparing data from different studies, and may indicate a further level of control for ligand binding in vivo

Expression, regulation and clinical significance of soluble and membrane CD14 receptors in pediatric inflammatory lung diseases

<p>Abstract</p> <p>Background</p> <p>Inflammatory lung diseases are a major morbidity factor in children. Therefore, novel strategies for early detection of inflammatory lung diseases are of high interest. Bacterial lipopolysaccharide (LPS) is recognized via Toll-like receptors and CD14. CD14 exists as a soluble (sCD14) and membrane-associated (mCD14) protein, present on the surface of leukocytes. Previous studies suggest sCD14 as potential marker for inflammatory diseases, but their potential role in pediatric lung diseases remained elusive. Therefore, we examined the expression, regulation and significance of sCD14 and mCD14 in pediatric lung diseases.</p> <p>Methods</p> <p>sCD14 levels were quantified in serum and bronchoalveolar lavage fluid (BALF) of children with infective (pneumonia, cystic fibrosis, CF) and non-infective (asthma) inflammatory lung diseases and healthy control subjects by ELISA. Membrane CD14 expression levels on monocytes in peripheral blood and on alveolar macrophages in BALF were quantified by flow cytometry. <it>In vitro </it>studies were performed to investigate which factors regulate sCD14 release and mCD14 expression.</p> <p>Results</p> <p>sCD14 serum levels were specifically increased in serum of children with pneumonia compared to CF, asthma and control subjects. <it>In vitro</it>, CpG induced the release of sCD14 levels in a protease-independent manner, whereas LPS-mediated mCD14 shedding was prevented by serine protease inhibition.</p> <p>Conclusions</p> <p>This study demonstrates for the first time the expression, regulation and clinical significance of soluble and membrane CD14 receptors in pediatric inflammatory lung diseases and suggests sCD14 as potential marker for pneumonia in children.</p

Expression, regulation and clinical significance of soluble and membrane CD14 receptors in pediatric inflammatory lung diseases

<p>Abstract</p> <p>Background</p> <p>Inflammatory lung diseases are a major morbidity factor in children. Therefore, novel strategies for early detection of inflammatory lung diseases are of high interest. Bacterial lipopolysaccharide (LPS) is recognized via Toll-like receptors and CD14. CD14 exists as a soluble (sCD14) and membrane-associated (mCD14) protein, present on the surface of leukocytes. Previous studies suggest sCD14 as potential marker for inflammatory diseases, but their potential role in pediatric lung diseases remained elusive. Therefore, we examined the expression, regulation and significance of sCD14 and mCD14 in pediatric lung diseases.</p> <p>Methods</p> <p>sCD14 levels were quantified in serum and bronchoalveolar lavage fluid (BALF) of children with infective (pneumonia, cystic fibrosis, CF) and non-infective (asthma) inflammatory lung diseases and healthy control subjects by ELISA. Membrane CD14 expression levels on monocytes in peripheral blood and on alveolar macrophages in BALF were quantified by flow cytometry. <it>In vitro </it>studies were performed to investigate which factors regulate sCD14 release and mCD14 expression.</p> <p>Results</p> <p>sCD14 serum levels were specifically increased in serum of children with pneumonia compared to CF, asthma and control subjects. <it>In vitro</it>, CpG induced the release of sCD14 levels in a protease-independent manner, whereas LPS-mediated mCD14 shedding was prevented by serine protease inhibition.</p> <p>Conclusions</p> <p>This study demonstrates for the first time the expression, regulation and clinical significance of soluble and membrane CD14 receptors in pediatric inflammatory lung diseases and suggests sCD14 as potential marker for pneumonia in children.</p

Model based analysis of real-time PCR data from DNA binding dye protocols

BACKGROUND: Reverse transcription followed by real-time PCR is widely used for quantification of specific mRNA, and with the use of double-stranded DNA binding dyes it is becoming a standard for microarray data validation. Despite the kinetic information generated by real-time PCR, most popular analysis methods assume constant amplification efficiency among samples, introducing strong biases when amplification efficiencies are not the same. RESULTS: We present here a new mathematical model based on the classic exponential description of the PCR, but modeling amplification efficiency as a sigmoidal function of the product yield. The model was validated with experimental results and used for the development of a new method for real-time PCR data analysis. This model based method for real-time PCR data analysis showed the best accuracy and precision compared with previous methods when used for quantification of in-silico generated and experimental real-time PCR results. Moreover, the method is suitable for the analyses of samples with similar or dissimilar amplification efficiency. CONCLUSION: The presented method showed the best accuracy and precision. Moreover, it does not depend on calibration curves, making it ideal for fully automated high-throughput applications

Synchrony of hand-foot coupled movements: is it attained by mutual feedback entrainment or by independent linkage of each limb to a common rhythm generator?

BACKGROUND: Synchrony of coupled oscillations of ipsilateral hand and foot may be achieved by controlling the interlimb phase difference through a crossed kinaesthetic feedback between the two limbs, or by an independent linkage of each limb cycle to a common clock signal. These alternative models may be experimentally challenged by comparing the behaviour of the two limbs when they oscillate following an external time giver, either alone or coupled together. RESULTS: Ten subjects oscillated their right hand and foot both alone and coupled (iso- or antidirectionally), paced by a metronome. Wrist and ankle angular position and Electromyograms (EMG) from the respective flexor and extensor muscles were recorded. Three phase delays were measured: i) the clk-mov delay, between the clock (metronome beat) and the oscillation peak; ii) the neur (neural) delay, between the clock and the motoneurone excitatory input, as inferred from the EMG onset; and iii) the mech (mechanical) delay between the EMG onset and the corresponding point of the limb oscillation. During uncoupled oscillations (0.4 Hz to 3.0 Hz), the mech delay increased from -7° to -111° (hand) and from -4° to -83° (foot). In contrast, the clk-mov delay remained constant and close to zero in either limb since a progressive advance of the motoneurone activation on the pacing beat (neur advance) compensated for the increasing mech delay. Adding an inertial load to either extremity induced a frequency dependent increase of the limb mechanical delay that could not be completely compensated by the increase of the neural phase advance, resulting in a frequency dependent increment of clk-mov delay of the hampered limb. When limb oscillations were iso- or antidirectionally coupled, either in the loaded or unloaded condition, the three delays did not significantly change with respect to values measured when limbs were moved separately. CONCLUSION: The absence of any significant effect of limb coupling on the measured delays suggests that during hand-foot oscillations, both iso- and antidirectionally coupled, each limb is synchronised to the common rhythm generator by a "private" position control, with no need for a crossed feedback interaction between limbs