Limitations In Terms Of Resolution Research Articles

Analytic Validation of Optical Genome Mapping in Hematological Malignancies.

Structural variations (SVs) play a key role in the pathogenicity of hematological malignancies. Standard-of-care (SOC) methods such as karyotyping and fluorescence in situ hybridization (FISH), which have been employed globally for the past three decades, have significant limitations in terms of resolution and the number of recurrent aberrations that can be simultaneously assessed, respectively. Next-generation sequencing (NGS)-based technologies are now widely used to detect clinically significant sequence variants but are limited in their ability to accurately detect SVs. Optical genome mapping (OGM) is an emerging technology enabling the genome-wide detection of all classes of SVs at a significantly higher resolution than karyotyping and FISH. OGM requires neither cultured cells nor amplification of DNA, addressing the limitations of culture and amplification biases. This study reports the clinical validation of OGM as a laboratory-developed test (LDT) according to stringent regulatory (CAP/CLIA) guidelines for genome-wide SV detection in different hematological malignancies. In total, 60 cases with hematological malignancies (of various subtypes), 18 controls, and 2 cancer cell lines were used for this study. Ultra-high-molecular-weight DNA was extracted from the samples, fluorescently labeled, and run on the Bionano Saphyr system. A total of 215 datasets, Inc.luding replicates, were generated, and analyzed successfully. Sample data were then analyzed using either disease-specific or pan-cancer-specific BED files to prioritize calls that are known to be diagnostically or prognostically relevant. Sensitivity, specificity, and reproducibility were 100%, 100%, and 96%, respectively. Following the validation, 14 cases and 10 controls were run and analyzed using OGM at three outside laboratories showing reproducibility of 96.4%. OGM found more clinically relevant SVs compared to SOC testing due to its ability to detect all classes of SVs at higher resolution. The results of this validation study demonstrate the superiority of OGM over traditional SOC methods for the detection of SVs for the accurate diagnosis of various hematological malignancies.

Diamond nitrogen-vacancy color-centered thermometer for integrated circuit application.

With the advancement of the chip industry, accurate temperature measurement and thermal management have become crucial. Traditional infrared temperature imaging has limitations in terms of resolution and accuracy. lnrecent years, quantum diamond nitrogen-vacancy centers have emerged as a promising option for temperature sensing, but separating temperature from magnetic field effects remains a challenge. This paper presents a numerical approach to decouple temperature and magnetic fields using an ensemble Hamiltonian in high-current density Integrated Circuit (IC) applications. The proposed method demonstrates a temperature sensitivity of 22.9 mK/Hz1/2 and the ability to perform scanning temperature imaging with a spatial resolution of 20 µm on a typical IC.

Impact of sea level changes on future wave conditions along the coasts of western Europe

Abstract. Wind waves and swells are major drivers of coastal environment changes and coastal hazards such as coastal flooding and erosion. Wave characteristics are sensitive to changes in water depth in shallow and intermediate waters. However, wave models used for historical simulations and projections typically do not account for sea level changes whether from tides, storm surges, or long-term sea level rise. In this study, the sensitivity of projected changes in wave characteristics to the sea level changes is investigated along the Atlantic European coastline. For this purpose, a global wave model is dynamically downscaled over the northeastern Atlantic for the 1970–2100 period under the SSP5–8.5 climate change scenario. Twin experiments are performed with or without the inclusion of hourly sea level variations from regional 3D ocean simulations in the regional wave model. The largest impact of sea level changes on waves is located on the wide continental shelf where shallow-water dynamics prevail, especially in macro-tidal areas. For instance, in the Bay of Mont-Saint-Michel in France, due to an average tidal range of 10 m, extreme historical wave heights were found to be up to 1 m higher (+30 %) when sea level variations are included. At the end of the 21st century, extreme significant wave heights are larger by up to +40 % (+60 cm), mainly due to the effect of tides and mean sea level rise. The estimates provided in this study only partially represent the processes responsible for the sea-level–wave non-linear interactions due to model limitations in terms of resolution and the processes included.

P456: A LONG READ SEQUENCING AND CRISPR-CAS9 BASED APPROACH FOR RAPID COPY-NUMBER ALTERATION AND STRUCTURAL VARIATION DETECTION IN HEMATOLOGIC MALIGNANCIES

Background: Current standard karyotyping methods are labor-intensive and have severe limitations in terms of resolution and duration to results. For hematologic malignancies, but not restricted to, this represents a relevant obstacle in translating molecular findings into time critical treatment decisions. Aims: To develop a sequencing based approach for rapid and scalable cytogenetics that can be applied to a wide spectrum of hematologic malignancies to detect clinically relevant molecular markers within 48hrs Methods: For copy number alteration detection, whole genome sequencing was performed on a GridION sequencer (Oxford Nanopore Technologies, ONT, using the SQK-LSK109 kit) in a cohort of 53 hematologic neoplasms, including acute myeloid leukemia (AML, n=18), chronic lymphocytic leukemia (CLL, n=8), multiple myeloma (MM, n=17) and pediatric acute lymphocytic leukemia (ALL, n=10) samples. For the identification of balanced alterations, transcriptome sequencing data were generated using ONT (SQK-DCS109 kit), and we applied an in-house developed analysis pipeline based on minimap2 alignment followed by Blast with an ensuing filtering algorithm based on orientation, inter-read distance and length filtering. For targeted sequencing of structural aberrations, we developed a CRISPR-Cas9 based tiling approach using sgRNA pools (Integrated DNA Technologies, IDT). This approach allows to cover AML relevant targets such as t(8;21), the KMT2A (MLL) breakpoint area, and the FLT3-ITD region as well as lymphoma relevant regions such as the immunoglobulin heavy chain locus (library preparation was performed using SQK-CS9109 kit). Results: For copy number profiling a median whole-genome coverage of 2.6 was reached (range 0.27-7 fold). ONT sequencing and conventional karyotyping approaches showed a high concordance with Pearson correlation coefficients >0.95 for copy number alteration comparisons between conventional cytogenetics and ONT sequencing results for all investigated disease entities. Regarding the detection of structural aberrations transcriptome sequencing and fusion gene analysis using the above described analysis and filtering pipeline we could e.g. reliably detect the t(9;22) translocation from the K562 cell line and the t(8;21) translocation from Kasumi-1. Additionally, using this method we were able to correctly identify the primary AML sample harboring a t(8;21) translocation that was among the n=12 AML patient samples. In addition to an RNA-based work-flow, we established a CRISPR-Cas9 based DNA enrichment approach for the detection of recurrent structural aberrations from specified genomic DNA loci. The enrichment of genomic regions of interest using sgRNA libraries led e.g. to a median coverage of 44 reads (range 5-136-fold) of the t(8;21) region-of-interest in primary AML samples, which allow the detection of the exact coordinates of the breakpoints. Summary/Conclusion: Long read based sequencing based copy number alteration and structural variation detection based on combination of different long read sequencing approaches like low-coverage whole genome sequencing, transcriptome sequencing and CRISPR-Cas9 based sequencing of genomic loci of interest represents a highly promising tool for high resolution and high speed cytogenetics that has the potential to overcome many limitations of conventional cytogenetics.

Intraspecies-Resolution Metabarcoding: automated primer design and a plant pathology case study

We present results demonstrating species-specificity and sub-species resolution by novel, automatically-designed metabarcoding primers for environmental DNA analysis. Conventional metabarcoding remains a cornerstone of rapid, high-throughput environmental DNA (eDNA) community analysis and biodiversity assessment. Standard barcodes such as 16S (prokaryotes) and ITS1 (fungi/oomycetes) have been instrumental in identifying the complex composition of communities using total eDNA. However, standard barcodes have limitations in terms of resolution and quantitation and, though genus-level identification can be reliable, species-level identification is often not possible. To overcome the limitations of resolution, we implemented extensions to the diagnostic primer design tool pdp (https://github.com/widdowquinn/find_differential_primers [https://github.com/widdowquinn/find_differential_primers]) that enable automated design of metabarcoding markers and corresponding primers that are (i) specific to a prescribed taxon at species level and (ii) capable of discriminating between members of the same species. This allows for rapid, high-throughput measurement of diversity below species level for a target organism. We aimed to survey geographical distribution and pathogen transfer of the widespread plant pathogenic bacterium Pectobacterium atrosepticum (Pba). This organism has considerable sub-species taxonomic structure identifiable using MLST and with whole-genome methods, but which is not accessible using standard barcodes. We designed metabarcoding primers (202bp) specific to Pba using `pdp`, and established that these have resolution comparable to eight-gene MLST, revealing sub species-level diversity within single fields, and on the same individual plant host.

Multiview tiling light sheet microscopy for 3D high-resolution live imaging.

Light-sheet or selective plane illumination microscopy (SPIM) is ideally suited for in toto imaging of living specimens at high temporal-spatial resolution. In SPIM, the light scattering that occurs during imaging of opaque specimens brings about limitations in terms of resolution and the imaging field of view. To ameliorate this shortcoming, the illumination beam can be engineered into a highly confined light sheet over a large field of view and multi-view imaging can be performed by applying multiple lenses combined with mechanical rotation of the sample. Here, we present a Multiview tiling SPIM (MT-SPIM) that combines the Multi-view SPIM (M-SPIM) with a confined, multi-tiled light sheet. The MT-SPIM provides high-resolution, robust and rotation-free imaging of living specimens. We applied the MT-SPIM to image nuclei and Myosin II from the cellular to subcellular spatial scale in early Drosophila embryogenesis. We show that the MT-SPIM improves the axial-resolution relative to the conventional M-SPIM by a factor of two. We further demonstrate that this axial resolution enhancement improves the automated segmentation of Myosin II distribution and of nuclear volumes and shapes.

Interferometric device for the in-process measurement of diameter variation in the manufacture of ultraprecise spheres

A novel device is presented which is designed for in-process measurements of the variation of the diameter of highly reflective spheres. Silicon spheres have been used for the new definition of the International System of Units (SI). Many spheres have to be processed, and the form of these objects, and thus the manufacturing process’s stability, needs to be controlled every day. Commercially available measurement equipment and even state-of-the-art spherical interferometers have reached their limits in terms of resolution, uncertainty, the complexity of their handling routines, measurement time and even financial investment. A novel setup has thus been designed after considering and selecting a special mechanical setup with a minimal measurement loop, stable optical sensors and a handling strategy which avoids collision and contact with the very valuable, superpolished spherical objects. Thus, the design minimizes the influence of the environment and reduces the measurement time at an equator with sub-nanometre resolution to 3 min. In addition, the analysis time is reduced to less than a minute.

Stepwise Optimized 3D Printing of Arbitrary 3D Structures at Millimeter Scale with High Precision Surface

Abstract3D printing based on additive manufacturing has attracted widespread attention in the fields of microbiology and microelectronics due to its advantages of waste reduction, arbitrary manufacturing, and rapid prototyping in potential applications. These techniques can create structures at the centimeter scale, however, there are some limitations in terms of resolution and geometric constraints. Here, a micro–nano 3D printing protocol based on additive manufacturing to achieve the 3D structure (3DS) not only possessing millimeter scale structural dimensions but also nanometer features are proposed. A theory is verified to assist the design and fabrication of the 3DS with millimeter scale and nanometer precision. The structures are predesigned and the scanning strategy is optimized before 3D printing to improve the manufacturing efficiency and precision. A customized 3DS with a height of 2.2 mm is obtained, which is a challenge for the conventional two‐photon polymerization fabrication. Furthermore, a 1.2 mm 3DS with inside scaffold and smooth surface is efficiently achieved within 2.7 h with a nanometer surface roughness by using the proposed stepwise optimized 3D printing process. This study offers a flexible and low‐cost technology to generate highly customizable, precisely controllable 3DS for potential applications in microelectronics and microdevices.

New Imaging Modalities in Otology.

Despite steady improvements in cross-sectional imaging of the ear, current technologies still have limitations in terms of resolution, diagnosis, functional assessment and safety. In this chapter, state-of-the-art imaging techniques in current clinical practice are presented including cone-beam computerized tomography, non-echo planar imaging magnetic resonance imaging, imaging for labyrinthine hydrops and imaging of the central auditory pathways. Potential future imaging modalities are also presented, including optical coherence tomography (OCT) and high-frequency ultrasound (HFUS) of the ear. These experimental modalities offer new opportunities for the assessment of ear structure and function. For example, middle ear structures can be visualized through the tympanic membrane, basilar membrane vibrations can be assessed through the round window and the passage of cochlear implants can be assessed in decalcified cochlear. Functional assessment of the middle ear using Doppler techniques are also discussed, including measurement of tympanic membrane and middle ear vibration amplitudes, visualization of dynamic changes, such as tensor tympani movements and movement of the tympanic membrane with breathing. These new modalities currently have limitations that preclude mainstream clinical use. For example, OCT is limited by the optical scattering of the thickened tympanic membrane and HFUS needs a coupling medium such as gel or fluid from the transducer to the imaged structure although it can visualize through thicker tissues. Nevertheless, further development of these novel techniques may provide an enhanced ability to assess the ear in conjunction with current technologies.

A new star tracker concept for satellite attitude determination based on a multi-purpose panoramic camera

This paper presents an innovative algorithm developed for attitude determination of a space platform. The algorithm exploits images taken from a multi-purpose panoramic camera equipped with hyper-hemispheric lens and used as star tracker. The sensor architecture is also original since state-of-the-art star trackers accurately image as many stars as possible within a narrow- or medium-size field-of-view, while the considered sensor observes an extremely large portion of the celestial sphere but its observation capabilities are limited by the features of the optical system. The proposed original approach combines algorithmic concepts, like template matching and point cloud registration, inherited from the computer vision and robotic research fields, to carry out star identification. The final aim is to provide a robust and reliable initial attitude solution (lost-in-space mode), with a satisfactory accuracy level in view of the multi-purpose functionality of the sensor and considering its limitations in terms of resolution and sensitivity. Performance evaluation is carried out within a simulation environment in which the panoramic camera operation is realistically reproduced, including perturbations in the imaged star pattern. Results show that the presented algorithm is able to estimate attitude with accuracy better than 1° with a success rate around 98% evaluated by densely covering the entire space of the parameters representing the camera pointing in the inertial space.

High-Throughput Immunogenetics for Clinical and Research Applications in Immunohematology: Potential and Challenges

Analysis and interpretation of Ig and TCR gene rearrangements in the conventional, low-throughput way have their limitations in terms of resolution, coverage, and biases. With the advent of high-throughput, next-generation sequencing (NGS) technologies, a deeper analysis of Ig and/or TCR (IG/TR) gene rearrangements is now within reach, which impacts on all main applications of IG/TR immunogenetic analysis. To bridge the generation gap from low- to high-throughput analysis, the EuroClonality-NGS Consortium has been formed, with the main objectives to develop, standardize, and validate the entire workflow of IG/TR NGS assays for 1) clonality assessment, 2) minimal residual disease detection, and 3) repertoire analysis. This concerns the preanalytical (sample preparation, target choice), analytical (amplification, NGS), and postanalytical (immunoinformatics) phases. Here we critically discuss pitfalls and challenges of IG/TR NGS methodology and its applications in hemato-oncology and immunology.

CLIP-ASAP-HSQC for fast and accurate extraction of one-bond couplings from isotropic and partially aligned molecules.

Fast measurement of heteronuclear one-bond couplings, a class of NMR parameters valuable for structure elucidation, is highly desirable, especially if samples undergo chemical reactions or dynamic processes are observed. Methods presented so far face severe limitations in terms of resolution, accessible bandwidth, and sensitivity. We present the CLean InPhase-Acceleration by Sharing Adjacent Polarization-HSQC (CLIP-ASAP-HSQC) pulse sequence that allows fast acquisition of spectra with clean inphase multiplets in about 25 s. The performance in terms of accurate extraction of one-bond couplings is demonstrated on three test samples including partially aligned molecules.

Spectral estimation-What is new? What is next?

Spectral estimation, and corresponding time-frequency representation for nonstationary signals, is a cornerstone in geophysical signal processing and interpretation. The last 10–15 years have seen the development of many new high-resolution decompositions that are often fundamentally different from Fourier and wavelet transforms. These conventional techniques, like the short-time Fourier transform and the continuous wavelet transform, show some limitations in terms of resolution (localization) due to the trade-off between time and frequency localizations and smearing due to the finite size of the time series of their template. Well-known techniques, like autoregressive methods and basis pursuit, and recently developed techniques, such as empirical mode decomposition and the synchrosqueezing transform, can achieve higher time-frequency localization due to reduced spectral smearing and leakage. We first review the theory of various established and novel techniques, pointing out their assumptions, adaptability, and expected time-frequency localization. We illustrate their performances on a provided collection of benchmark signals, including a laughing voice, a volcano tremor, a microseismic event, and a global earthquake, with the intention to provide a fair comparison of the pros and cons of each method. Finally, their outcomes are discussed and possible avenues for improvements are proposed.

A high-throughput X-ray-based method for measurements of relative wood density from unprepared increment cores from Picea abies

We have described a high-throughput method for ring width and relative wood density analysis of unprepared increment cores based on conventional computer tomography scanners (CT scanners) used in health care. The method was developed for analysis of 5-mm increment cores collected in forest genetic and tree breeding trials. For these studies, the relative differences between individuals are more important than the exact value. The method is highly efficient in terms of quantitative data collection from large number of samples. There are, however, limitations in terms of resolution and accuracy on very small volumes of wood that must be considered when using the method to measure individual growth rings or increment cores thinner than 5 mm. However, in studies comprising large number of samples, the use of cost-efficient CT scanning on the major part of the material, combined with more laborious high-resolution scanning on sub-samples can be a good combination for efficient determination of radial variation in growth rate and wood density.

Studying Electrocatalytic Activity Using Scanning Electrochemical Microscopy

Scanning electrochemical microscopy is an excellent tool for studying electrocatalytic reactions at the nanometer scale, although it has not been much utilized for this purpose. So far, most of the electrocatalysis studies reported using SECM are mainly focused on the relationship between bulk chemical composition and electrochemical activity. This article presents some examples of electrocatalytic reactions studied by SECM and reviews the progress on the use of SECM to study electrocatalytic reactions, paying special attention to studying the surface structure effect on nanoparticles and translating of single crystal electrode results into electrocatalysts based on shape-controlled nanoparticles. SECM currently does have some limitations in terms of resolution of the interrogated nanometric domains. Hence, in the future, it will be necessary to develop new combined electrochemical probe techniques and nanoprobes to overcome said limitations and perhaps approach probing electrocatalysis at the surface of a single nanoparticle.

Evaluation of Version-7 TRMM Multi-Satellite Precipitation Analysis Product during the Beijing Extreme Heavy Rainfall Event of 21 July 2012

The latest Version-7 (V7) Tropical Rainfall Measuring Mission (TRMM) Multi-satellite Precipitation Analysis (TMPA) products were released by the National Aeronautics and Space Administration (NASA) in December of 2012. Their performance on different climatology, locations, and precipitation types is of great interest to the satellite-based precipitation community. This paper presents a study of TMPA precipitation products (3B42RT and 3B42V7) for an extreme precipitation event in Beijing and its adjacent regions (from 00:00 UTC 21 July 2012 to 00:00 UTC 22 July 2012). Measurements from a dense rain gauge network were used as the ground truth to evaluate the latest TMPA products. Results are summarized as follows. Compared to rain gauge measurements, both 3B42RT and 3B42V7 generally captured the rainfall spatial and temporal pattern, having a moderate spatial correlation coefficient (CC, 0.6) and high CC values (0.88) over the broader Hebei, Beijing and Tianjin (HBT) regions, but the rainfall peak is 6 h ahead of gauge observations. Overall, 3B42RT showed higher estimation than 3B42V7 over both HBT and Beijing. At the storm center, both 3B42RT and 3B42V7 presented a relatively large deviation from the temporal variation of rainfall and underestimated the storm by 29.02% and 36.07%, respectively. The current study suggests that the latest TMPA products still have limitations in terms of resolution and accuracy, especially for this type of extreme event within a latitude area on the edge of coverage of TRMM precipitation radar and microwave imager. Therefore, TMPA users should be cautious when 3B42RT and 3B42V7 are used to model, monitor, and forecast both flooding hazards in the Beijing urban area and landslides in the mountainous west and north of Beijing.

Performance evaluation of radar and satellite rainfalls for Typhoon Morakot over Taiwan: Are remote-sensing products ready for gauge denial scenario of extreme events?

summary This study evaluated rainfall estimates from ground radar network and four satellite algorithms with a relatively dense rain gauge network over Taiwan Island for the 2009 extreme Typhoon Morakot at various spatiotemporal scales (from 0.04 to 0.25 and hourly to event total accumulation). The results show that all the remote-sensing products underestimate the rainfall as compared to the rain gauge measurements, in an order of radar (� 18%), 3B42RT (� 19%), PERSIANN-CCS (28%), 3B42V6 (� 36%), and CMORPH (� 61%). The ground radar estimates are also most correlated with gauge measurements, having a correlation coefficient (CC) of 0.81 (0.82) at 0.04 (0.25) spatial resolution. For satellite products, CMORPH has the best spatial correlation (0.70) but largely underestimates the total rainfall accumulation. Compared to microwave ingested algorithms, the IR-dominant algorithms provide a better estimation of the total rainfall accumulation but poorly resolve the temporal evolution of the warm cloud typhoon, especially for a large overestimation at the early storm stage. This study suggests that the best performance comes from the ground radar estimates that could be used as an alternative in case of the gauge denial. However, the current satellite rainfall products still have limitations in terms of resolution and accuracy, especially for this type of extreme typhoon.

A saline tracer test monitored via both surface and cross-borehole electrical resistivity tomography: Comparison of time-lapse results

In contrast to traditional field investigation techniques in hydrogeology, geophysical methods are relatively non-invasive, cost effective and can be performed with a higher spatial sampling. The most commonly applied technique in hydrogeophysics is electrical resistivity tomography (ERT), both from the ground surface and in cross-borehole configurations. To infer reliable results from such a hydrogeophysical application, however, the uncertainty related to the data inversion has to be taken into account and specific attention must be paid to the experimental set-up and design, especially when the main target of the study is a quantitative estimation of some relevant hydrological quantity. The sensitivity and resolving power of ERT depend on the type of acquisition methodology; operating from the ground surface only, for example, could lead to severe limitations in terms of resolution, thus limiting the quantitative utilisation from a hydrogeological perspective. In this work, we present the results of a saline tracer test experiment performed in the saturated zone at the water works facility at Valdobbiadene (Treviso, North-East Italy), where an alluvial phreatic aquifer is heavily exploited for irrigation and drinking water supply. The experiment was monitored by time-lapse ERT acquisitions, using both surface and cross-borehole configurations. We compared the results of the two approaches and conclude that, in general, ERT has excellent imaging capabilities for saline tracer tests, however, significant limitations are inherent in the use of surface electrode configurations only. (C) 2011 Elsevier B.V. All rights reserved.

State-of-the-Art Characterization for 65 nm CMOS Processes and Beyond

Characterization of advanced CMOS devices and control of front- end of line (FEOL) processes require state-of-the-art analytical techniques and specific measurement protocols. Dynamic Secondary Ion Mass Spectrometry (D-SIMS) is well suited for in-depth quantitative analysis of both bulk- and dopant concentrations in GexSi1-x binary alloys. A similar D-SIMS protocol -in combination with Auger Electron Spectroscopy (AES) - allows monitoring nickel-silicide phase transitions upon silicidation of (doped) Ni Totally Silicided (TOSI) gates. Combination of Low-Energy Electron X-ray Emission Spectroscopy (LEXES) and high-precision DSIMS provides accurate control of dose, junction depth, and wafer uniformity for Ultra-Shallow Junctions (USJ) at high analytical throughput, opening the way to USJ metrology. Similar techniques (LEXES, SIMS) are employed to control wafer uniformity of deposited NiPt metal films as Source/Drain (S/D) contact metal, to monitor NixSi1-x phase transformations and Pt-redistribution upon silicidation, and to determine root causes of defectivity at the device level (micro- AES). The given examples demonstrate the importance of having available (novel) state-of-the-art characterization techniques - pushing the analytical limits in terms of resolution, sensitivity, and accuracy while simultaneously increasing throughput and reproducibility- to support process development, improve yield, and to speed up yield learning of advanced CMOS processes.

X-Ray Diffraction as a Probe for Elastic Strain: Micro- and Nanoscale Investigation of Thin Metal Films

ABSTRACTIn the past years the concept of measuring strain by x-rays has changed significantly. The combination of 3rd generation synchrotron sources, advanced focusing techniques and large area detectors has made it possible to probe volumes smaller than a cubic micron. This devolopment has made it possible to probe microstrains directly without having to rely on highly sophisticated models to evaluate peak broadening effects. This paper will provide a review of the state of art of local strain measurements by x-rays, discuss their limitations, provide an outlook of where the field may be going within the next years and address the most important issues to be solved. Examples will be given for the current limits in terms of resolution in time, space, strain and intensity.