Application of self-quenched JH consensus primers for real-time quantitative PCR of IGH gene to minimal residual disease evaluation in multiple myeloma.

Monitoring multiple myeloma patients for relapse requires sensitive methods to measure minimal residual disease and to establish a more precise prognosis. The present study aimed to standardize a real-time quantitative polymerase chain reaction (PCR) test for the IgH gene with a JH consensus self-quenched fluorescence reverse primer and a VDJH or DJH allele-specific sense primer (self-quenched PCR). This method was compared with allele-specific real-time quantitative PCR test for the IgH gene using a TaqMan probe and a JH consensus primer (TaqMan PCR). We studied nine multiple myeloma patients from the Spanish group treated with the MM2000 therapeutic protocol. Self-quenched PCR demonstrated sensitivity of >or=10(-4) or 16 genomes in most cases, efficiency was 1.71 to 2.14, and intra-assay and interassay reproducibilities were 1.18 and 0.75%, respectively. Sensitivity, efficiency, and residual disease detection were similar with both PCR methods. TaqMan PCR failed in one case because of a mutation in the JH primer binding site, and self-quenched PCR worked well in this case. In conclusion, self-quenched PCR is a sensitive and reproducible method for quantifying residual disease in multiple myeloma patients; it yields similar results to TaqMan PCR and may be more effective than the latter when somatic mutations are present in the JH intronic primer binding site.

Minimal residual disease monitoring in multiple myeloma: a comparison between allelic-specific oligonucleotide real-time quantitative polymerase chain reaction and flow cytometry.

BACKGROUND AND OBJECTIVES: Minimal residual disease (MRD) studies are useful in multiple myeloma (MM). However, the definition of the best technique and clinical utility are still unresolved issues. The aim of this study was to analyze and compare the clinical utility of MRD studies in MM with two different techniques: allelic-specific oligonucleotide real-time quantitative PCR (ASO-RQ-PCR), and flow cytometry (FCM). DESIGN AND METHODS: Bone marrow samples from 32 MM patients who had achieved complete response after transplantation were evaluated by ASO-RQ-PCR, using TaqMan technology, and multiparametric FCM. RESULTS: ASO-RQ-PCR was only applicable in 75% of patients for a variety of technical reasons, while FCM was applicable in up to 90%. Therefore, simultaneous PCR/FCM analysis was possible in only 24 patients. The number of residual tumor cells identified by both techniques was very similar (mean=0.29%, range=0.001-1.61%, correlation coefficient=0.861). However, RQ-PCR was able to detect residual myelomatous cells in 17 patients while FCM only did so in 11; thus, 6 cases were FCM negative but PCR positive, all of them displaying a very low number of clonal cells (median=0.014%, range=0.001-0.11). Using an MRD threshold of 0.01% (10(-4)) two risk groups with significantly different progression-free survival could be identified by either PCR (34 vs. 15m, p=0.04) or FCM (27 vs. 10m, p=0.05). INTERPRETATION AND CONCLUSIONS: Although MRD evaluation by ASO-RQ-PCR is slightly more sensitive and specific than FCM, it is applicable in a lower proportion of MM patients and is more time-consuming, while both techniques provide similar prognostic information.

Incomplete DJH rearrangements as a novel tumor target for minimal residual disease quantitation in multiple myeloma using real-time PCR.

The hypervariable regions of immunoglobulin heavy-chain (IgH) rearrangements provide a specific tumor marker in multiple myeloma (MM). Recently, real-time PCR assays have been developed in order to quantify the number of tumor cells after treatment. However, these strategies are hampered by the presence of somatic hypermutation (SH) in VDJH rearrangements from multiple myeloma (MM) patients, which causes mismatches between primers and/or probes and the target, leading to a nonaccurate quantification of tumor cells. Our group has recently described a 60% incidence of incomplete DJH rearrangements in MM patients, with no or very low rates of SH. In this study, we compare the efficiency of a real-time PCR approach for the analysis of both complete and incomplete IgH rearrangements in eight MM patients using only three JH consensus probes. We were able to design an allele-specific oligonucleotide for both the complete and incomplete rearrangement in all patients. DJH rearrangements fulfilled the criteria of effectiveness for real-time PCR in all samples (ie no unspecific amplification, detection of less than 10 tumor cells within 10(5) polyclonal background and correlation coefficients of standard curves higher than 0.98). By contrast, only three out of eight VDJH rearrangements fulfilled these criteria. Further analyses showed that the remaining five VDJH rearrangements carried three or more somatic mutations in the probe and primer sites, leading to a dramatic decrease in the melting temperature. These results support the use of incomplete DJH rearrangements instead of complete somatically mutated VDJH rearrangements for investigation of minimal residual disease in multiple myeloma.

Why are egg fertilization rates not correlated with hatching success in sea turtles?

[EN] The relative number of developing eggs is directly affected by fertilization rate, and unfertile eggs may indirectly negatively affect development of viable eggs within the nest. Thus, the number of viable eggs at laying should influence hatching success. We have studied both parameters in a nesting population of loggerhead turtles from Boavista Island (Republic of Cabo Verde). Fertility was estimated based on eggs excavated from nests within the first 96 hours after deposition. Our results confirm a high egg fertilization rate for the species, which exceeded an average of 94% fertility (95% confidence limits: 91.9 and 96.2%, N=43 nests).

Generic wind estimation and compensation based on residual generators and higher-order sliding mode schemes

This master thesis proposes a solution to the approach problem in case of unknown severe microburst wind shear for a fixed-wing aircraft, accounting for both longitudinal and lateral dynamics. The adaptive controller design for wind rejection is also addressed, exploiting the wind estimation provided by suitable estimators. It is able to successfully complete the final approach phase even in presence of wind shear, and at the same time aerodynamic envelope protection is retained. The adaptive controller for wind compensation has been designed by a backstepping approach and feedback linearization for time-varying systems. The wind shear components have been estimated by higher-order sliding mode schemes. At the end of this work the results are provided, an autonomous final approach in presence of microburst is discussed, performances are analyzed, and estimation of the microburst characteristics from telemetry data is examined.

Residual Stresses and Fatigue Behavior of Welded Structural Members, HR-94, 1967

A research project entitled "Residual Stresses and Fatigue Behavior of Welded Structural Members" was conducted at the Structural Research Laboratory of the Engineering Research Institute at Iowa State University under the sponsorship of the Iowa State Highway Commission. The objective of the project was to study experimentally the fatigue behavior of flange plates in welded beam sections as influenced by different residual stress distributions which are caused by different sizes of welds.

Medición del desempeño : retorno sobre inversión, ROI; ingreso residual, IR; valor económico agregado, EVA; análisis comparado

El artículo quiere a partir de un recuento crítico de los análisis tradicionales del ROI y del IR dar una mirada analítica a estos indicadores comparándolos entre sí, mirando su alcance e importancia reales, igualmente sus problemas conceptuales y limitaciones. Al final se introduce el EVA, como medida y filosofía, mirando lo novedoso de la misma y los aspectos comunescon el ROI y el IR.

Sistemas de produção de milho sob adubação nitrogenada e plantas de cobertura do solo

The no-tillage system is the predominant model in the agricultural scenario of southern Brazil. Thus, the use of cover crops is significant due to the addition of biomass to protect the soil surface, and contribute to the cycling and/or fixing of nutrients, and in particular nitrogen (N) with liberation for the subsequent culture. Among the cool season species, it was found predominant use of oat to obtain straw to system. Though large quantities input of residue is not the preferred species to precede the corn, cereal with relevant importance in the Paraná Southwest region. It was aimed to evaluate the productivity capacity of corn in no-tillage, in the absence or presence of nitrogen fertilization, on waste of winter cover crops on soil and climatic conditions of the Paraná Southwest region. The installation of no-tillage was held in 2010 in the experimental area belonging to UTFPR, Campus Dois Vizinhos, on a Red Latosol. For the present study, we used data relating to three agricultural years (2012/2013, 2013/2014 and 2014/2015). The experimental design was randomized block design with split plots with three replications. The main plots consisted of systems composed by cover crops (black oat, ryegrass, rye, turnip, vetch, white lupine, aot+vetch consortium and oat+vetch+turnip), preceding corn. In the subplots were used two doses of nitrogen fertilization (0 and 180 kg ha N) coverage in maize.The biggest coverage rates occurred in the consortium with 95% at 62 days after sowing. The residual effect of 180 kg ha cool season plants following year. The residual effect of 180 kg ha systems, reduced in 21% the C/N ratio of poaceae. The common vetch accumulated 32 kg N per ton of MS added. The oat and rye keeps more than 50% waste to the land cover, after 120 days, while the ryegrass and vetch provide low soil protection. Consortium oat+vetch+turnip, vetch and white lupine, released the largest amounts of N, between 52 and 59 kg ha brassica and consortia positively influencing the diameter and length of cobs, number of kernels per row and, total number of grains per ear of corn, in the absence of mineral N. The weight of a thousand grains was increased by 12.4% by the addition of 180 kg ha increase in productivity of grain by the addition of 180 kg ha N, was 2.1 Mg ha 5.6 Mg ha 6.4 Mg ha components when cultivated on vetch. Systems containing fabaceae, brassica and consortium oat+vetch+turnip, predating the corn, in the absence of mineral N, provided similar grain yelds inrelation to the systems with the addition of 180 kg ha Keywords: Cover crops. No-tillage. Grain yield. Zea mays - 1 -1 N, increased 4.8% coverage rate in the of N in corn/cover crops -1 -1 . Fabaceae, -1 N mineral. The average N, in relation to dose 0 kg ha corn kernels on fabaceae, brassica and consortium oat+vetch+turnip, and poaceae the grains in succession. The consortium added amount between 4.0 the DM in the years of study. There was no effect of mineral N rate for corn yield components when cultivated on vetch. Systems containing fabaceae, brassica and consortium oat+vetch+turnip, predating the corn, in the absence of mineral N, provided similar grain yelds inrelation to the systems with the addition of 180 kg ha-1 N.

Characterization of Non-linear Polymer Properties to Predict Process Induced Warpage and Residual Stress of Electronic Packages

Nonlinear thermo-mechanical properties of advanced polymers are crucial to accurate prediction of the process induced warpage and residual stress of electronics packages. The Fiber Bragg grating (FBG) sensor based method is advanced and implemented to determine temperature and time dependent nonlinear properties. The FBG sensor is embedded in the center of the cylindrical specimen, which deforms together with the specimen. The strains of the specimen at different loading conditions are monitored by the FBG sensor. Two main sources of the warpage are considered: curing induced warpage and coefficient of thermal expansion (CTE) mismatch induced warpage. The effective chemical shrinkage and the equilibrium modulus are needed for the curing induced warpage prediction. Considering various polymeric materials used in microelectronic packages, unique curing setups and procedures are developed for elastomers (extremely low modulus, medium viscosity, room temperature curing), underfill materials (medium modulus, low viscosity, high temperature curing), and epoxy molding compound (EMC: high modulus, high viscosity, high temperature pressure curing), most notably, (1) zero-constraint mold for elastomers; (2) a two-stage curing procedure for underfill materials and (3) an air-cylinder based novel setup for EMC. For the CTE mismatch induced warpage, the temperature dependent CTE and the comprehensive viscoelastic properties are measured. The cured cylindrical specimen with a FBG sensor embedded in the center is further used for viscoelastic property measurements. A uni-axial compressive loading is applied to the specimen to measure the time dependent Young’s modulus. The test is repeated from room temperature to the reflow temperature to capture the time-temperature dependent Young’s modulus. A separate high pressure system is developed for the bulk modulus measurement. The time temperature dependent bulk modulus is measured at the same temperatures as the Young’s modulus. The master curve of the Young’s modulus and bulk modulus of the EMC is created and a single set of the shift factors is determined from the time temperature superposition. The supplementary experiments are conducted to verify the validity of the assumptions associated with the linear viscoelasticity. The measured time-temperature dependent properties are further verified by a shadow moiré and Twyman/Green test.

Altura do pasto e adubação nitrogenada sobre a produção de forragem e eficiência no uso de nutrientes em sistema de integração lavoura-pecuária

The proper use of management strategies, such as grazing intensity and nitrogen fertilization are primordial to the success of integrated crop-livestock system. Several studies have demonstrated the influence of grazing intensity and nitrogen fertilization on dynamics of forage production and nutrient cycling. However, most this researches studying these strategies in isolation and little is known about the interaction of these factors in the management of an integrated crop-livestock system. In this context, the aim of this study is to determine the best management strategy involving sward height and nitrogen fertilization, permitting greater forage production and improved efficiency in the use of nitrogen soil by a black oat ‘BRS 139’ plus ryegrass ‘Barjumbo’ pasture in integrated crop-livestock system. The experiment was realized in Abelardo Luz – SC, in an area of 14 ha, where has been conducted an experiment in long term with integrated crop-livestock system under no-tillage since 2012. The experimental design is a randomized block design with three replications in a factorial design (2x2), the first factor was the grazing intensity (high and low), characterized by two sward height management (10 and 25 cm), and the second included the time factor application of N in the system: N applied on pasture (N-pasture) and N applied on the culture of grain (N-grain), at dose of 200 kg N ha stocking and variable stocking rate. The previous crop to pasture was corn. The nitrogen fertilization of pasture increased tiller density, forage density, participation of ryegrass ‘Barjumbo’ and percentage of ryegrass leaves in forage mass. Forage mass was less at low sward height on average, however the percentage of ryegrass ‘Barjumbo’ and rye leaves was greater and dead material was lower in this treatment. With nitrogen fertilization of pasture it was possible to double the amount of forage accumulated in periods with further development of ryegrass, furthermore, the total production of DM was increased in 38.4% and the shoot N concentration in 28.6%. When the nitrogen fertilization is applied in pasture, it is possible to keep black oat ‘BRS 139’ plus ryegrass ‘Barjumbo’ pasture with an average sward height of 11 cm. The residual effect of N applied at corn was not sufficient to meet the nutritional needs of pasture and the forage production was affected by periods with N deficiency, while a single application of 200 kg N ha was sufficient to meet the N requirements throughout the forage accumulation period. The black oat ‘BRS 139’ plus ryegrass ‘Barjumbo’ pasture is efficient in use and recovery of the nitrogen applied in both treatments of sward height.

Aplicação foliar de molibdênio em adubos verdes e o desempenho do feijoeiro em sucessão

The knowledge of molybdenum application in legumes on the availability of N, by BNF, increased enzymatic activity and the residual effect caused on crops growth and yield can contribute to the greater scientific understanding involved in green manure processes. The aim of this study was to evaluate the Mo application and the N from Crotalaria juncea and Canavalia ensiformis green manures on common bean performance. Were conducted field experiments for the crops succession system (green manures - common bean) and laboratory essays for the enzymatic activities. Green manure production was installed in a factorial arrangement 2 x 4, with two green manure legumes species, sunnhemp (Crotalaria juncea) and jack beans (Canavalia ensiformis), and four Mo doses (0, 40, 80, 120 g ha-1) in the form of sodium molybdate (Na2MoO4), foliar applied, in a randomized block design with four replicates. For succession crop (common bean) additional treatment was added, beans grown without any fertilization, following the same experimental design from the previous crop. The dry matter decomposition and the N mineralization of green manure were monitored through collection of residues over time, by using the litter bags method. In laboratory were carried out tests of nitrate reductase activity in green manures and common beans at 90 and 66 days after sowing, respectively. The sunnhemp responded linearly positively to the application of Mo as the dry matter and N accumulation. While the jack beans presented a negative quadratic response for dry matter and there was no adjustment of regression models to N. The jack beans showed a higher decomposition rate and N mineralization compared to sunnhemp. The half lives for decomposing 50% of dry matter on the soil was 123 and 104 days to sunnhemp and jack beans, respectively, and 50% of N present in the residues was mineralized at 93 and 85 days. In common bean, differed from the control for number of pods the dose of 40 g ha-1 of Mo in both species of green manures and the dose 80 g ha-1 of Mo in jack beans. For number of grains only in sunnhemp on the dose of 40 g ha-1 of Mo differ from the control. The nitrate reductase activity was influenced by developmental stage of green manure species. In common bean, the activity of nitrate reductase was up to three times higher than the dose 0 g ha-1 of Mo compared to treatment with application of Mo in both species. There was no effect of Mo doses or species of green manure on common bean yield.

Fertilization strategies for Sea Bass Dicentrarchus labrax (Linnaeus, 1758): effects of pre-incubation and duration of egg receptivity in seawater

Studying gamete biology can provide important information about a species fertilization strategy as well as their reproductive ecology. Currently, there is a lack of knowledge about how long sea bass Dicentrarchus labrax eggs can remain viable after being activated in seawater. The objectives of this study were to understand the effects of pre-incubation of fresh and overripe sea bass eggs in seawater and to determine the duration of egg receptivity. Pooled eggs (fresh and overripe) from four females were pre-incubated in seawater for 0 min (control), 0.5 min, 1 min, 3 min, 10 min and 30 min and then fertilized by pooled sperm from four males. The fresh eggs had a higher fertilization success than overripe eggs. Our results revealed a significant effect of pre-incubation time for both the fresh (P < 0.01) and overripe eggs (P < 0.01). Fertilization success of eggs significantly declined for both these treatments after 3 min of pre-incubation, which clearly indicates that sea bass eggs are able to be fertilized by sperm for up to 3 min after release into seawater. This study has particular importance for understanding fertilization strategies, reproductive potential, as well as reproductive ecology of sea bass.