Evaluation of a PCR probe capture assay for the detection of Toxoplasma gondii: incorporation of uracil N-glycosylase for contamination control

Journal ArticleToxoplasma gondii is a cyst-forming parasite of clinical relevance in humans primarily because of the neurologic abnormalities it can cause. In some clinical circumstances, it is desirable to detect the pathogen directly. We modified a commercially available Toxoplasma polymerase chain reaction (PCR) probe capture assay by incorporating uracil N-glycosylase (UNG) to prevent carryover amplicon contamination. In addition, UNG inactivation and DNA denaturation were accomplished chemically to simplify the DNA hybridization to the capture probe. The incorporation of UNG effectively eliminated carryover contamination; the probe capture assay showed a log increase in detection sensitivity compared with standard agarose gel electrophoresis. To assess sensitivity and possible inhibition of amplification, different sample types were spiked with Toxoplasma organisms. After DNA extraction and PCR amplification, a sensitivity of 2 tachyzoites for the assay was determined in buffered saline, cerebrospinal fluid (CSF), serum, and amniotic fluid; 20 tachyzoites for whole blood; and 200 tachyzoites for brain tissue. An additional 20 human serum and CSF samples submitted for Toxoplasma serologic testing were run by the PCR method. Of these, only an IgM-positive CSF sample was PCR positive. The Toxoplasma PCR probe capture assay showed good sensitivity and was not substantially inhibited by several different clinically relevant samples

Comparison of four enzyme immunoassays with a western blot assay for the determination of type-specific antibodies to herpes simplex virus

Journal ArticleMost current enzyme immunoassays (EIAs) differentiate inadequately between types 1 and 2 herpes simplex virus (HSV) antibodies since significant crossreactivity exists. We compared 4 IgG type-specific EIAs using a Western blot assay for resolution of discrepant results. The Diamedix had sensitivities of 100% for types 1 and 2 but specificities of only 71% and 61%, respectively. The cross-reactivity rate was 82% in positive samples tested. For HSV types 1 and 2, the Zeus sensitivities were 92% and 98%, respectively; specificities were 72% and 79%, respectively; the cross-reactivity rate was 54%. For HSV types 1 and 2, the Wampole sensitivities were 98% and 95%, respectively; specificities were 68% and 85%, respectively; the cross-reactivity rate was 47%. For HSV types 1 and 2, the Meridian sensitivities were 98% and 90%, respectively; specificities were 96% and 100%, respectively; no cross-reactivity was found between positive samples tested. While the Diamedix, Zeus, and Wampole assays showed good sensitivity, they lacked type specificity. The Meridian EIA offers the highest specificity along with no observed cross-reactivity. This EIA may be an easier, reliable alternative to Western blot for the determination of HSV type-specific antibodies

Determination of cytokine responses using a multiplexed fluorescent microsphere immunoassay

Journal ArticleWe used a multiplexed fluorescent microsphere immunoassay to develop a sandwich capture assay to assess simultaneously the production of thymus helper (TH) 1- and TH2-type cytokines in tissue culture supernatant obtained from stimulated peripheral blood mononuclear cells. The assay then was used to assess the cytokine production of patients with hyperimmunoglobulinemia E syndrome and in cord blood from neonates. The multiplexed assay has a reportable range of less than 10 to 50,000 pg/mL. For linearity and recovery studies, R2 values for the 6 cytokines ranged from 0.988 to 0.999 for samples spiked with known concentrations of recombinant cytokine standards and for patient samples. The assay showed good specificity, with little cross-reactivity between cytokines. Results from supernatants of Staphylococcus aureus-stimulated peripheral blood mononuclear cells obtained from 6 patients with hyperimmunoglobulinemia E syndrome showed significantly less interferon (IFN)-gamma production than cells from healthy control subjects. Cord blood cells from neonates produced significantly less interleukin 12 and IFN-gamma than cells from adults in group B streptococci-stimulated mononuclear cells. The fluorescent multiplexed microsphere immunoassay can be used to quantitate multiple cytokines from 1 sample and should be useful for further understanding of the cytokine role in disease

Stenosis of the Branches of the Neopulmonary Artery after the Arterial Switch Operation: a Cardiac Magnetic Resonance Imaging Study

Background : The neonatal arterial switch operation (ASO) is now the standard of care for children born with transposition of the great arteries. Stenosis of the neopulmonary artery on long‑term follow up is a known complication. Methods : We performed a retrospective analysis of eleven patients who underwent a cardiac magnetic resonance imaging (MRI) due to echocardiographic evidence suggestive of stenosis of the neopulmonary artery or its branches (mean estimated Doppler gradient 48 mmHg, min 30 mmHg, max 70 mmHg). A comprehensive evaluation of anatomy and perfusion was done by cardiac MRI. Results : The branches of the neopulmonary artery (neo PA) showed decreased caliber in three patients unilaterally and in two patients, bilaterally. Magnetic resonance (MR) perfusion studies showed concomitant decreased flow, with discrepancy between the two lungs of 35/65% or worse, only in the three patients with unilateral obstruction, by two different MR perfusion methods. Conclusions : Cardiac MR can be used as a comprehensive non‑invasive imaging technique to diagnose stenosis of the branches of the neopulmonary after the ASO, allowing evaluation of anatomy and function of the neoPA, its branches, and the differential perfusion to each lung, thus facilitating clinical decision making

Analyzing the Spectral Energy Cascade in Turbulent Channel Flow

An analysis of the spectral turbulent kinetic energy budget in a fully developed turbulent plane channel flow is performed. Direct numerical simulation data are evaluated at friction Reynolds numbers Reτ of 180 and 1000. The analysis is focused on the influence of the Reynolds number on the spectral cascade of energy and the corresponding energy cascade in physical space in the presence of inhomogeneity and anisotropy. The turbulent kinetic energy distribution is compared for both Reynolds numbers, and the relevant characteristics of the energy transfer process in a wall-bounded turbulent flow are described. Differences in energy cascade are noted between the Reynolds number at both low and high wavenumbers. The results of the analysis are further assessed with a comparison to an earlier study of spectral energy transfer at Reτ= 180

Compensatory growth in oceanic loggerhead sea turtles: response to a stochastic environment

Compensatory growth (CG, accelerated growth that may occur when an organism that has grown at a reduced rate as a result of suboptimal environmental conditions is exposed to better conditions) is considered an adaptation to variable en vironments. Although documented thoroughly under captive conditions, CG has rarely been studied in wild populations. In their first years of life, oceanic-stage loggerhead sea turtles (Caretta caretta) have relatively little control over their geographic position or movements and thus have an extremely stochastic lifestyle with great variation in food availability and temperature. This environmental variation results in variable growth rates. We evaluate somatic growth functions of oceanic-stage loggerheads from the eastern Atlantic based on skeletochronology that allowed us to assign age and cohort to each individual. We demonstrate CG in these turtles based on three different analytical approaches: changes in coefficients of variation in size-at-age, generalized additive model regression analyses of somatic growth, and linear regression of age-specific growth rates. As a result of CG, variation in size-at-age in these juvenile loggerheads is substantially reduced. Thus, size is a better predictor of age than expected based on variation in growth rates. CG decreases with age, apparently as loggerheads gain greater control over their movements. In addition, we have evaluated for the first time in wild sea turtles the time-dependent nature of somatic growth by distinguishing among age, year, and cohort effects using a mixed longitudinal sampling design with assigned-age individuals. Age and year had significant effects on growth rates, but there was no significant cohort effect. Our results address critical gaps in knowledge of the demog raphy of this endangered species.info:eu-repo/semantics/publishedVersio

Fast spin exchange between two distant quantum dots

The Heisenberg exchange interaction between neighboring quantum dots allows precise voltage control over spin dynamics, due to the ability to precisely control the overlap of orbital wavefunctions by gate electrodes. This allows the study of fundamental electronic phenomena and finds applications in quantum information processing. Although spin-based quantum circuits based on short-range exchange interactions are possible, the development of scalable, longer-range coupling schemes constitutes a critical challenge within the spin-qubit community. Approaches based on capacitative coupling and cavity-mediated interactions effectively couple spin qubits to the charge degree of freedom, making them susceptible to electrically-induced decoherence. The alternative is to extend the range of the Heisenberg exchange interaction by means of a quantum mediator. Here, we show that a multielectron quantum dot with 50-100 electrons serves as an excellent mediator, preserving speed and coherence of the resulting spin-spin coupling while providing several functionalities that are of practical importance. These include speed (mediated two-qubit rates up to several gigahertz), distance (of order of a micrometer), voltage control, possibility of sweet spot operation (reducing susceptibility to charge noise), and reversal of the interaction sign (useful for dynamical decoupling from noise).Comment: 6 pages including 4 figures, plus 8 supplementary pages including 5 supplementary figure

Transatlantic developmental migrations of loggerhead sea turtles demonstrated by mtDNA sequence analysis

Molecular markers based on mitochondrial (mt) DNA control region se quences were used to test the hypothesis that juvenile loggerhead sea turtles (Caretta caretta) in pelagic habitats of the eastern Atlantic are derived from nesting populations in the western Atlantic. We compared mtDNA haplotypes from 131 pelagic juvenile turtles (79 from the Azores and 52 from Madeira) to mtDNA haplotypes observed in major nesting colonies of the Atlantic Ocean and Mediterranean Sea. A subset of 121 pelagic samples (92%) contained haplotypes that match mtDNA sequences observed in nesting colonies. Maximum likelihood analyses (UCON, SHADRACQ) estimate that 100% of these pelagic juveniles are from the nesting populations in the southeastern United States and adjacent Yucatan Peninsula, Mexico. Estimated contributions from nesting populations in south Florida (0.71, 0.72), northern Florida to North Carolina (0.19, 0.17), and Quintana Roo, Mexico (0.11, 0.10) are consistent with the relative size of these nesting aggregates. No contribution was detected from nesting colonies in the Mediterranean (Greece) or South Atlantic (Brazil), although samples sizes are insufficient to exclude these locations with finality. The link between west Atlantic nesting colonies and east Atlantic feeding grounds provides a more complete scientific basis for assessing the impact of subadult mortality in oceanic fisheries. Demographic models for loggerhead turtles in the western Atlantic can now be improved by incorporating growth and mortality data from juvenile turtles in pelagic habitats. These data demonstrate that the appropriate scale for loggerhead turtle conservation efforts is vastly larger than the current scale of management plans based on political boundaries.info:eu-repo/semantics/publishedVersio

Chromosome differentiation patterns during cichlid fish evolution

<p>Abstract</p> <p>Background</p> <p>Cichlid fishes have been the subject of increasing scientific interest because of their rapid adaptive radiation which has led to an extensive ecological diversity and their enormous importance to tropical and subtropical aquaculture. To increase our understanding of chromosome evolution among cichlid species, karyotypes of one Asian, 22 African, and 30 South American cichlid species were investigated, and chromosomal data of the family was reviewed.</p> <p>Results</p> <p>Although there is extensive variation in the karyotypes of cichlid fishes (from 2n = 32 to 2n = 60 chromosomes), the modal chromosome number for South American species was 2n = 48 and the modal number for the African ones was 2n = 44. The only Asian species analyzed, <it>Etroplus maculatus</it>, was observed to have 46 chromosomes. The presence of one or two macro B chromosomes was detected in two African species. The cytogenetic mapping of 18S ribosomal RNA (18S rRNA) gene revealed a variable number of clusters among species varying from two to six.</p> <p>Conclusions</p> <p>The karyotype diversification of cichlids seems to have occurred through several chromosomal rearrangements involving fissions, fusions and inversions. It was possible to identify karyotype markers for the subfamilies Pseudocrenilabrinae (African) and Cichlinae (American). The karyotype analyses did not clarify the phylogenetic relationship among the Cichlinae tribes. On the other hand, the two major groups of Pseudocrenilabrinae (tilapiine and haplochromine) were clearly discriminated based on the characteristics of their karyotypes. The cytogenetic mapping of 18S ribosomal RNA (18S rRNA) gene did not follow the chromosome diversification in the family. The dynamic evolution of the repeated units of rRNA genes generates patterns of chromosomal distribution that do not help follows the phylogenetic relationships among taxa. The presence of B chromosomes in cichlids is of particular interest because they may not be represented in the reference genome sequences currently being obtained.</p