Erratum: Available Energy of Trapped Electrons and Its Relation to Turbulent Transport [Phys. Rev. Lett. 128, 175001 (2022)].

This corrects the article DOI: 10.1103/PhysRevLett.128.175001.

Enhanced Transport at High Plasma Pressure and Subthreshold Kinetic Ballooning Modes in Wendelstein 7-X.

High-performance fusion plasmas, requiring high pressure ß, are not well understood in stellarator-type experiments. Here, the effect of ß on ion-temperature-gradient-driven (ITG) turbulence is studied in Wendelstein 7-X (W7-X), showing that subdominant kinetic ballooning modes (KBMs) are unstable well below the ideal MHD threshold and get strongly excited in the turbulence. By zonal-flow erosion, these subthreshold KBMs (stKBMs) affect ITG saturation and enable higher heat fluxes. Controlling stKBMs will be essential to allow W7-X and future stellarators to achieve maximum performance.

Available Energy of Trapped Electrons and Its Relation to Turbulent Transport.

Any collisionless plasma possesses some "available energy" (AE), defined as that part of the thermal energy that can be converted into instabilities and turbulence. Here, we present a calculation of the AE carried by magnetically trapped electrons in a flux tube of collisionless plasma. The AE is compared with nonlinear simulations of the energy flux resulting from collisionless turbulence driven by trapped-electron modes in various magnetic geometries. The numerical calculation of AE is rapid and shows a strong correlation with the simulated energy fluxes, which can be expressed as a power law and understood in terms of a simple model.

Turbulence Mechanisms of Enhanced Performance Stellarator Plasmas.

We theoretically assess two mechanisms thought to be responsible for the enhanced performance observed in plasma discharges of the Wendelstein 7-X stellarator experiment fueled by pellet injection. The effects of the ambipolar radial electric field and the electron density peaking on the turbulent ion heat transport are separately evaluated using large-scale gyrokinetic simulations. The essential role of the stellarator magnetic geometry is demonstrated, by comparison with a tokamak.

Controlling turbulence in present and future stellarators.

Turbulence is widely expected to limit the confinement and, thus, the overall performance of modern neoclassically optimized stellarators. We employ novel petaflop-scale gyrokinetic simulations to predict the distribution of turbulence fluctuations and the related transport scaling on entire stellarator magnetic surfaces and reveal striking differences to tokamaks. Using a stochastic global-search optimization method, we derive the first turbulence-optimized stellarator configuration stemming from an existing quasiomnigenous design.

Resilience of quasi-isodynamic stellarators against trapped-particle instabilities.

It is shown that in perfectly quasi-isodynamic stellarators, trapped particles with a bounce frequency much higher than the frequency of the instability are stabilizing in the electrostatic and collisionless limit. The collisionless trapped-particle instability is therefore stable as well as the ordinary electron-density-gradient-driven trapped-electron mode. This result follows from the energy balance of electrostatic instabilities and is thus independent of all other details of the magnetic geometry.

A multi-site study using high-resolution HLA genotyping by next generation sequencing.

The high degree of polymorphism at human leukocyte antigen (HLA) class I and class II loci makes high-resolution HLA typing challenging. Current typing methods, including Sanger sequencing, yield ambiguous typing results because of incomplete genomic coverage and inability to set phase for HLA allele determination. The 454 Life Sciences Genome Sequencer (GS FLX) next generation sequencing system coupled with conexio atf software can provide very high-resolution HLA genotyping. High-throughput genotyping can be achieved by use of primers with multiplex identifier (MID) tags to allow pooling of the amplicons generated from different individuals prior to sequencing. We have conducted a double-blind study in which eight laboratory sites performed amplicon sequencing using GS FLX standard chemistry and genotyped the same 20 samples for HLA-A, -B, -C, DPB1, DQA1, DQB1, DRB1, DRB3, DRB4, and DRB5 (DRB3/4/5) in a single sequencing run. The average sequence read length was 250 base pairs and the average number of sequence reads per amplicon was 672, providing confidence in the allele assignments. Of the 1280 genotypes considered, assignment was possible in 95% of the cases. Failure to assign genotypes was the result of researcher procedural error or the presence of a novel allele rather than a failure of sequencing technology. Concordance with known genotypes, in cases where assignment was possible, ranged from 95.3% to 99.4% for the eight sites, with overall concordance of 97.2%. We conclude that clonal pyrosequencing using the GS FLX platform and CONEXIO ATF software allows reliable identification of HLA genotypes at high resolution.

Overcoming methodical limits of standard RHD genotyping by next-generation sequencing.

BACKGROUND AND OBJECTIVES: Molecular variations of the RHD gene may result in the reduced expression of the D antigen and altered Rh phenotypes. In many occasions, they cannot be typed reliably by standard serological methods. Sequence-based typing is the gold standard to determine rare and unknown RHD genotypes. For this pilot study, sequence-based typing by standard Sanger sequencing was compared to a newly established next-generation sequencing approach based on pyrosequencing. MATERIALS AND METHODS: Twenty-six DNA samples were selected after primary serological testing exhibiting a weak reaction in Rh phenotype. Parallel sequence analysis of the complete coding sequence including adjacent intronic sequences allowed a comparison of the methodical potency in mutation detection of Sanger with next-generation sequencing. RESULTS: Sanger sequencing revealed 39 RHD polymorphisms in 21 of 26 samples in the RHD coding region, while pyrosequencing detected all but two alterations resulting in a concordance rate of 94·9% and clearly revealed a heterozygous compound mutation in one sample with RHDψ and Weak D type 4 alleles. The resolution of cis/trans linkage of polymorphisms and exact characterization of a 37 bp duplication was achieved by next-generation sequencing. CONCLUSION: Our data suggest that next-generation sequencing offers a new development for high-throughput and clonal sequencing for molecular RHD genotyping. However, further attempts in the methodical set-up have to be undertaken prior to validation and introduction as a routine service.

Characterisation of two novel HLA alleles, HLA-Cw*0429 and HLA-DRB3*0223.

We report here the novel human leukocyte antigens (HLA)-Cw*0429 and HLA-DRB3*0223 alleles identified during routine cord blood characterisation by sequence-based typing.

Introduction of a real-time-based blood-group genotyping approach.

BACKGROUND AND OBJECTIVE: Genotyping may be applied for rare blood group polymorphisms in a high-throughput mode as well for the molecular determination of blood groups due to unclear serological results. MATERIAL AND METHODS: We developed and validated a DNA typing method for the determination of KEL1/2, JK1/2, FY1/2, FY0, MNS1/2, MNS3/4, DO1/2, CO1/2 and LU1/2 alleles using a melting curve analysis downstream from a fully automated DNA extraction. All assays were validated in terms of specificity, sensitivity, assay variability and robustness. The usability was proven by a batch of 200 blood samples with partially known phenotype. RESULTS: Assays for all blood groups were within the range of specificity (100%), assay variability and robustness (coefficient of variance < 3%). Genotypes of 200 random platelet donors were fully consistent with existing phenotype data. The obtained genotype distribution is in complete concordance with existing data for the European population underlined by a complete absence of CO2 homozygous donors and the FY0 allele among the cohort. CONCLUSION: We introduce an approach for blood group genotyping of particular samples or gene loci in glass capillary format and for medium-throughput analysis in 96/384-well format. The advantages of this real-time polymerase chain reaction method are its automation potential, the flexibility regarding hardware and the rapid cycling time.

The full length HLA-G1 and no other alternative form of HLA-G is expressed at the cell surface of transfected cells.

In contrast to HLA class Ia, the HLA-G class Ib transcripts can be alternativeley spliced to yield several isoforms including four potentially membrane-bound variants, namely HLA-G1, -G2, -G3 and G4. It is so far unclear whether each of these splice variants lacking one or two external domains is properly translated and expressed at the cell surface. We used targeted Enhanced Green Fluorescence Protein (EGFP)-HLA-G fusion cDNA to track HLA-G isoform expression in living murine (L-human beta2m) and human (JAR) transiently transfected cells. It was demonstrated that the four HLA-G1, -G2, -G3, and -G4 isoforms were translated in these transfectants by the means of (i) Western blotting analysis, using an anti-EGFP mAb; (ii) intracellular double labeling flow cytometry analysis, using the EGFP natural fluorescence and phycoerythrin-labeled HCA2 anti-HLA-G mAb; and (iii) immunocytochemistry on isolated acetone fixed transfectants with the use of different anti-HLA-G mAbs. Cell surface flow cytometry analysis using the HCA2 mAb revealed that only the HLA-G1 isoform was expressed as a membrane-bound protein. Two color confocal microscopy performed on fixed, permeabilized cells further showed that the EGFP green fluorescence co-localized with anti-calnexin rhodamine fluorescence in the four HLA-G isoform transfectants but only in HLA-G1 transfectant was the green EGFP fluorescence also detectable at the outer part of the cells, suggesting that the HLA-G2, -G3, and G4 were retained in the endoplasmic reticulum. Such intracellular retention of the three shorter forms of HLA-G suggest that they may play a role in regulating cell surface expression either of the full length HLA-G1 form or of HLA-E.

Plasma protein carbonyl concentration is not enhanced by chronic intake of high-protein diets in adult rats.

We tested the hypothesis of whether high dietary protein intake is linked to oxidative stress as measured by the concentration of reactive carbonyl residues in plasma proteins. Three groups of male Wistar rats ( approximately 230 g, n = 10) were fed either 15% (15C), 30% (30C), or 60% (60C) casein diets over a period of 18 weeks. For comparison, a vitamin E deficient diet (60C-E) based on diet 60C was given to an additional group to provoke oxidative stress. Concentrations of alpha-tocopherol in plasma and of reactive carbonyl residues in total plasma proteins were measured by high performance liquid chromatography using fluorescence and by diode array detection after 2,4-dinitrophenylhydrazine reaction, respectively. After 1 week the concentration of reactive carbonyl residues in plasma proteins was found to be significantly (P < 0.05) higher in the 60C and 60C-E groups ( approximately 2.7 nmol/mg protein) compared with the 15C and 30C groups ( approximately 1.7 nmol/mg protein). After 14 weeks the 15C (3.4 +/- 1.2 nmol/mg protein) and 60C-E groups (3.9 +/- 1.7 nmol/mg protein) showed a significantly increased concentration of reactive carbonyl residues in plasma protein compared with the 30C and 60C groups (2.5 +/- 1.0 nmol/mg protein; 2.6 +/- 0.8 nmol/mg protein). As expected, chronic vitamin E deficiency (60C-E) resulted in significantly decreased alpha-tocopherol concentrations (3.91 +/- 2.42 micromol/mL vs. 31.3 +/- 4.8 micromol/mL) and a higher concentration of reactive carbonyl residues in plasma proteins. These results do not support the hypothesis that a chronic intake of high-protein diets leads to oxidative stress in adult rats. However, in the non-adapted state (1 week) a high protein intake contributes to oxidative modifications of protein-bound amino acid residues.

Feeding resistant starch affects fecal and cecal microflora and short-chain fatty acids in rats.

The effects of different forms of resistant potato starch (RS) on the major microbial population groups and short-chain fatty acids (SCFA) in the cecum and feces of rats were studied over a 5-mo feeding period. Thirty 8-wk-old male Wistar rats, averaging 210 g initial body weight, were adapted for 7 d to a balanced basal diet containing 60% waxy maize starch devoid of any RS. On d 8, three groups of 10 rats each were fed diets containing the following forms of starch: 1) rapidly digestible waxy maize starch (basal diet), 2) a mixture of 83.3% waxy maize starch and 16.7% native granular potato starch (RS 1), or 3) a mixture of 33.3% waxy maize starch and 66.7% modified potato starch (RS 2). The final RS content in RS 1 and RS 2 was 10%. Fecal samples were collected at d 8 and 1, 3, and 5 mo after the start of the experiment. Cecal contents were taken after 5 mo. The colony counts of microbial groups did not vary with time in the control or the RS 1 group (P > .05). Only the number of Bacteroides/fusobacteria decreased between mo 1 and 5 in rats fed RS 1 (P < .05). The RS 2 diet led to a significant increase in total culturable bacteria, lactobacilli, streptococci, and enterobacteria between mo 1 and 5. The RS 1 and RS 2 diets stimulated the growth of bifidobacteria. Cecal numbers of lactobacilli, streptococci, and enterobacteria were higher in rats fed RS 2 than in rats fed RS 1 or control diet (P < .05). Lactobacillus cellobiosus occurred only in rats fed RS 1 or RS 2. Acetate increased in mo 3 compared with d 8 in all groups (P < .05). The fecal and cecal SCFA displayed higher concentrations of acetate and propionate and a higher molar proportion of propionate in RS 2 than in RS 1 or control rats (P < .05). Stimulation of bifidobacteria, lactobacilli, and SCFA may be useful for the suppression of pathogenic organisms in the colon.

Rapid identification of compound mutations in patients with Philadelphia-positive leukaemias by long-range next generation sequencing.

An emerging problem in patients with Philadelphia (Ph)-positive leukaemias is the occurrence of cells with multiple mutations in the BCR-ABL1 tyrosine kinase domain (TKD) associated with high resistance to different tyrosine kinase inhibitors. Rapid and sensitive detection of leukaemic subclones carrying such changes, referred to as compound mutations, is therefore of increasing clinical relevance. However, current diagnostic methods including next generation sequencing (NGS) of short fragments do not optimally meet these requirements. We have therefore established a long-range (LR) NGS approach permitting massively parallel sequencing of the entire TKD length of 933bp in a single read using 454 sequencing with the GS FLX+ instrument (454 Life Sciences). By testing a series of individual and consecutive specimens derived from six patients with chronic myeloid leukaemia, we demonstrate that long-range NGS analysis permits sensitive identification of mutations and their assignment to the same or to separate subclones. This approach also facilitates readily interpretable documentation of insertions and deletions in the entire BCR-ABL1 TKD. The long-range NGS findings were reevaluated by an independent technical approach in select cases. Polymerase chain reaction (PCR) amplicons of the BCR-ABL1 TKD derived from individual specimens were subcloned into pGEM®-T plasmids, and >100 individual clones were subjected to analysis by Sanger sequencing. The NGS results were confirmed, thus documenting the reliability of the new technology. Long-range NGS analysis therefore provides an economic approach to the identification of compound mutations and other genetic alterations in the entire BCR-ABL1 TKD, and represents an important advancement of the diagnostic armamentarium for rapid assessment of impending resistant disease.

[Alpha-amylase inhibitors and soluble dietary fiber in rye: partial purification and effect on postprandial glycemia]. / alpha-Amylase-Inhibitoren und lösliche Ballaststoffe aus Roggen: Partielle Reinigung und Einfluss auf die postprandiale Glykämie.

The protein inhibitor of the alpha-amylase (D-type) and the soluble arabinoxylan of rye (Var. Clou) were isolated from flour and bran, respectively. The isolation of the alpha-amylase inhibitor involves the extraction of rye flour in aqueous CaCl2-solution (2 x 10(-3) M containing the hemicellulase preparation Veron HE (2 g/100 g flour), dialysis and lyophilization (preparation I) and further fractionation with ammonsulfate, using the fraction 20-50% for isolation (preparation II). The arabinoxylan isolation is carried out using extraction of rye bran in 80% ethanol (80 degrees C), centrifugation, aqueous extraction of the sediment, dialysis and lyophilization (preparation I). The further purification using the precipitate of the fraction 20-50% leads to preparation II. The alpha-amylase inhibitor preparation II and the arabinoxylan preparation II were applied in a diet containing wheat starch and casein and fed to diabetic and healthy rats (Levis and Wistar). The postprandial increase of glucose was determined. It was detected that the postprandial increase of glucose is influenced neither by the alpha-amylase inhibitor nor by the soluble arabinoxylan in comparison to the control experiments. However, the alpha-amylase inhibitor of wheat significantly decreases the postprandial increase of glucose. The application of a test meal with alpha-amylase inhibitor of rye to health and diabetic of type-II-volunteers showed no variation of the blood glucose values. The reduction of the increase of glucose by the soluble beta-glucan of oat cannot be confirmed for the soluble arabinoxylan of rye. We conclude that the effect of the alpha-amylase inhibitor as well as the soluble pentosan or glucan has to be examined for each cereal species.

[Absorption of triglycine, diglycine, glycine or equimolar mixtures of diglycine and glycine in the perfused small intestine of rats]. / Untersuchungen zur Absorption von Triglyzin, Diglyzin, Glyzin sowie äquimolaren Mischungen von Diglyzin und Glyzin am perfundierten Dünndarm der Ratte.

1. After a wash-out period of 20 min with Krebs-Henseleit-buffer a 20 cm segment of proximal or distal small intestine of nonanaesthesized rats got a bolus infusion of 3 ml either triglycine, diglycine, glycine or an equimolar mixture of diglycine and glycine, respectively, in a concentration range of 20 to 1000 mmol/l glycine equivalents. With 9 ml of a peptide and amino acid free solution the gut was perfused in a single-pass perfusion and the whole fluid recovered was investigated by quantitative thin layer chromatography for triglycine, diglycine and glycine. 2. In the concentration range up to 170 mmol/l glycine equivalents the nitrogen absorption is independent of the substrates perfused. In the highest concentration range an additional increased disappearance of triglycine could be found. This change cannot be observed to the same extent for diglycine, glycine or the mixture of both. In contrast, in the highest concentration range the competition between glycine and diglycine results in a decrease of absorption. 3. The proximal or distal nitrogen absorption of all substrates is similar. 4. In the perfusate the peptide and its splitting products were investigated. 5. Based on the discrepancy between the disappearance of triglycine and the appearance of its splitting products it is assumed. 1. that the transport of intact triglycine is of quantitative importance in the highest concentration range and 2. that in the lower concentration range membrane digestion followed the transport of the splitting products prevails. 6. Extending Ugoley's model for dipeptides three general variations for membrane digestion of tripeptides are proposed. One of this is likely for the triglycine absorption.

Postprandial thermogenesis of rats with glutamate induced obesity in relation to energy intake.

Postprandial thermogenesis was estimated in 4-month-old male rats with glutamate induced obesity after being fed with 300, 450 and 600 kJ/kg0.75 of a pellet diet, respectively by indirect calorimetry in computer-controlled open circuit metabolic cages over 8 h. After an intake of 600 kJ/kg0.75 (above the maintenance energy requirement) postprandial thermogenesis was significantly reduced in the obese animals to about 40% of control rats (12.0 versus 31.5 kJ/kg0.75 x 8h). It is concluded that the glutamate obese rat can be accepted as an animal model with impaired facultative thermogenesis, mainly caused by a reduction of sympathetic adrenergic activity.

Relations between enzyme activities connected with energy metabolism and parameters of food energy utilization in young and adult rats. Part 1. Studies on dietary effects on weight, body composition and energy gain.

Weaned male Wistar rats were fed ad libitum besides standard pellets a normal-protein (NP), a low-protein (LP) and a high-fat (HF) diet for different periods of time. In comparison to rats on the NP diet the body weight of the LP group was significantly lower after seven weeks feeding, not after a duration of about forty weeks. No remarkable diet-related differences were detected by determination of the mean body composition and of the relative gross energy content of the body neither after seven nor forty weeks. Total gross energy intake was practically identical both for the NP and LP rats irrespective on the duration of the feeding period. A significantly lower quotient of body energy gain/total gross energy intake under the protein-restrictive LP diet after seven weeks suggests the expression of an energy dissipation obviously no longer visible after a long time feeding period.

[Nutritional-physiological studies on acetylated fractions of protein from Vicia faba]. / Ernährungsphysiologische Untersuchungen an acetylierten Vicia-faba-Proteinfraktionen.

The nutritional-physiological value of an easily soluble and of a difficulty soluble fraction of protein from Vicia faba (showing a degree of acetylation of 0%, 43% and 94%, respectively) was characterized by determining the amino-acid content, the enzymatic amino-acid availability and the nitrogen balance in the rat. The results from both the amino-acid analysis and the animal experiments on the biological value evidenced that the nutritional-physiological value of the easily soluble fraction of protein from Vicia faba is significantly lower, which is in particular attributable to its lower content of sulphur-containing amino acids. Acetylation did not reduce the nutritional-physiological value of the easily soluble fraction of protein from Vicia faba, and impaired that of the difficulty soluble fraction but slightly. From the viewpoint of nutritional physiology, there are no objections against the use of these protein fractions as food additives.