Turing patterns on rotating spiral growing domains.

We investigate the emergence of Turing patterns in a system growing as a rotating spiral in two dimensions, utilizing the photosensitivity of the chlorine dioxide-iodine-malonic acid (CDIMA) reaction to control the growth process. We observe the formation of single and multiple (double and triple) stationary spiral patterns as well as transitional patterns. From numerical simulations of the Lengyel-Epstein model with an additional term to account for the effects of illumination on the reaction, we analyze the relationship between the final morphologies and the radial and angular growth velocities, identify conditions conducive to the formation of transitional structures, examine the importance of the size of the initial nucleation site in determining the spiral's multiplicity, and evaluate the stability and robustness of these Turing patterns. Our results indicate how inclusion of rotational degrees of freedom in the growth process may lead to the formation of a diverse new class of patterns in chemical and biological systems.

Insight into the role of copper-based materials against the coronaviruses MHV-3, a model for SARS-CoV-2, during the COVID-19 pandemic.

Coating high-touch surfaces with inorganic agents, such as metals, appears to be a promising long-term disinfection strategy. However, there is a lack of studies exploring the effectiveness of copper-based products against viruses. In this study, we evaluated the cytotoxicity and virucidal effectiveness of products and materials containing copper against mouse hepatitis virus (MHV-3), a surrogate model for SARS-CoV-2. The results demonstrate that pure CuO and Cu possess activity against the enveloped virus at very low concentrations, ranging from 0.001 to 0.1% (w/v). A greater virucidal efficacy of CuO was found for nanoparticles, which showed activity even against viruses that are more resistant to disinfection such as feline calicivirus (FCV). Most of the evaluated products, with concentrations of Cu or CuO between 0.003 and 15% (w/v), were effective against MHV-3. Cryomicroscopy images of an MHV-3 sample exposed to a CuO-containing surface showed extensive damage to the viral capsid, presumably due to the direct or indirect action of copper ions.

Prediction Techniques on FPGA for Latency Reduction on Tactile Internet.

Tactile Internet (TI) is a new internet paradigm that enables sending touch interaction information and other stimuli, which will lead to new human-to-machine applications. However, TI applications require very low latency between devices, as the system's latency can result from the communication channel, processing power of local devices, and the complexity of the data processing techniques, among others. Therefore, this work proposes using dedicated hardware-based reconfigurable computing to reduce the latency of prediction techniques applied to TI. Finally, we demonstrate that prediction techniques developed on field-programmable gate array (FPGA) can minimize the impacts caused by delays and loss of information. To validate our proposal, we present a comparison between software and hardware implementations and analyze synthesis results regarding hardware area occupation, throughput, and power consumption. Furthermore, comparisons with state-of-the-art works are presented, showing a significant reduction in power consumption of ≈1300× and reaching speedup rates of up to ≈52×.

A New Level II Oncoplastic Technique for Inferior Pole Defects: The Three-Petal Glandular Reconstruction (3-PR).

INTRODUCTION: Oncoplastic surgery (OPS) allows wide excisions and accurate tumor resection and reduces breast deformities by immediate reconstruction of large defects. Superior pedicled mammaplasties allow excellent results in large breasts. Conversely, loco-regional flaps are the standard of care in small-to-medium breasts. However, both techniques show limitations in case of large resections of the lower pole, resulting in skin retraction and downward deviation of nipple and areola. We present a new technique for inferior pole reconstruction to overcome these limitations. It is called "the three-petal reconstruction" (3-PR). METHODS: Between September 2016 and May 2019, ten patients with invasive breast cancer of the lower pole underwent breast conservation and 3-PR. RESULTS: The 3-PR was uneventful in all patients. No major or minor complications were recorded. Patient and surgeon evaluations scored as good to excellent in all cases. Surveillance examinations in the follow-up did not reveal calcifications nor any findings of suspicion within the reconstructed area. CONCLUSIONS: In case of very large defect of lower pole, the 3-PR reveals to be an easy, fast, reproducible method for inferior pole reconstruction. It can represent a niche between therapeutic mammaplasty and perforator flaps, and it could be added to existing available options for tailored reconstruction. LEVEL OF EVIDENCE III: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

M1 Polarized Macrophages Persist in Skin of Post-Bariatric Patients after 2 Years.

BACKGROUND: Obesity is considered a condition of systemic chronic inflammation. Under this condition, adipose tissue macrophages switch from an M2 (anti-inflammatory) activation pattern to an M1 (proinflammatory) activation pattern. OBJECTIVE: The study aimed to verify the profile of skin macrophage activation after bariatric surgery as well as the role of MMP-1 in extracellular tissue remodeling. METHODS: This is a prospective, controlled and comparative study with 20 individuals split into two groups according to their skin condition: post-bariatric and eutrophic patients. Histological and morphometric analyses based on hematoxylin-eosin, picrosirius red (collagen), orcein (elastic fiber systems), and alcian blue (mast cells)-stained sections and immunohistochemical analysis (CD68, iNOS, and mannose receptor) for macrophages and metalloproteinase-1 were performed. RESULTS: Post-bariatric skin showed an increase in inflammation, angiogenesis, CD68, M1 macrophages (P< 0.001), and mast cells (P< 0.01); a decrease in M2 macrophages (P< 0.01); and a significant decrease in the collagen fiber network (P< 0.001). MMP-1 was increased in the papillary dermis of post-bariatric skin and decreased in the epidermis compared to eutrophic skin (P< 0.05). CONCLUSION: This study shows that post-bariatric skin maintains inflammatory characteristics for two years. Mast cells and M1 macrophages maintain and enhance the remodeling of the dermal extracellular matrix initiated during obesity in part due to the presence of MMP-1 in the papillary dermis. EBM LEVEL IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Fully Parallel Implementation of Otsu Automatic Image Thresholding Algorithm on FPGA.

This work proposes a high-throughput implementation of the Otsu automatic image thresholding algorithm on Field Programmable Gate Array (FPGA), aiming to process high-resolution images in real-time. The Otsu method is a widely used global thresholding algorithm to define an optimal threshold between two classes. However, this technique has a high computational cost, making it difficult to use in real-time applications. Thus, this paper proposes a hardware design exploiting parallelization to optimize the system's processing time. The implementation details and an analysis of the synthesis results concerning the hardware area occupation, throughput, and dynamic power consumption, are presented. Results have shown that the proposed hardware achieved a high speedup compared to similar works in the literature.

Turing patterns modulation by chemical gradient in isothermal and non-isothermal conditions.

The modulation of Turing patterns through Dirichlet boundary conditions has been studied through the isothermal and non-isothermal versions of a Brusselator-like model in a small-size domain reactor. We considered the Minkowski functional and the rate of entropy production to characterize the morphological aspects of the patterns and to indicate transitions of spatial states. We find that boundary conditions can induce the spatial symmetry breaking of Turing patterns when they are defined around the equilibrium points of a homogeneous dynamical system. As a result, two different Turing patterns can emerge in a reactor under an imposed gradient of chemicals that contains the equivalent concentration of the equilibrium points at some point in the boundary.

A rare case of severe myositis as paraneoplastic syndrome on breast cancer.

BACKGROUND: Dermatomyositis and polymyositis are both types of idiopathic inflammatory myositis characterized by inflammation and weakness of proximal skeletal muscles and skin rash. CASE: A 49-year-old Caucasian woman recently diagnosed with breast cancer classified as T1N2M0, stage IIIA, presenting skin rash associated with heliotrope and Gottron's papules. In addition, there was a progression to a severe reduction in proximal muscle strength with severe dysphagia. The initial treatment was conducted, and the patient recovered from all symptoms and followed adjuvant cancer management. TREATMENT: At first, high dose of corticosteroid was administered as pulse therapy, and a radical mastectomy was indicated due to the severe symptoms of the paraneoplastic syndrome. Then chemotherapy and radiotherapy were applied, and oral corticoid associated with immunosupressive drug was administered for dermatomyositis control. DISCUSSION: The association between myositis and an increased risk of cancer has been demonstrated over the years. This patient has a high probability of dermatomyositis diagnosis. The initial treatment with high dose of glucocorticoids may result in an improvement of muscle lesions. Second-line treatment with azathioprine, methotrexate, or cyclophosphamide may be required for aggressive disease. Removal of the cancer induces improvement of paraneoplastic syndrome. CONCLUSION: Dermatomyositis can be a clinical manifestation of a paraneoplastic syndrome in patients with breast cancer. It is a rare diagnosis, and there is little evidence to guide treatment until now. It is possible to control the evolution of dermatomyositis with high doses of glucocorticoids in almost all cases; however, in severe cases of paraneoplastic syndrome, cancer treatment should start immediately.

Tailoring bisphosphonate-doped titanium films to optimally couple cellular responses and antibacterial activity for biomedical applications.

Titanium (Ti) is widely utilized as an implant material; nonetheless, its integration with bone tissue faces limitations due to a patient's comorbidities. To address this challenge, we employed a strategic approach involving the growth of thin films by spin-coating and surface functionalization with etidronate (ETI), alendronate (ALE), and risedronate (RIS). Our methodology involved coating of Ti cp IV disks with thin films of TiO2, hydroxyapatite (HA), and their combinations (1:1 and 1:2 v/v), followed by surface functionalization with ETI, ALE, and RIS. Bisphosphonate-doped films were evaluated in terms of surface morphology and physical-chemical properties by techniques such as electron microscopy, confocal microscopy, and x-ray photoelectron spectroscopy. The antibacterial potential of bisphosphonates alone or functionalized onto the Ti surface was tested against Staphylococcus aureus biofilms. Primary human bone mesenchymal stem cells were used to determine in vitro cell metabolism and mineralization. Although RIS alone did not demonstrate any antibacterial effect as verified by minimum inhibitory concentration assay, when Ti surfaces were functionalized with RIS, partial inhibition of Staphylococcus aureus growth was noted, probably because of the physical-chemical surface properties. Furthermore, samples comprising TiO2/HA (1:1 and 1:2 v/v) showcased an enhancement in the metabolism of nondifferentiated cells and can potentially enhance the differentiation of osteoblastic precursors. All samples demonstrated cell viability higher than 80%. Addition of hydroxyapatite and presence of bisphosphonates increase the metabolic activity and the mineralization of human bone mesenchymal cells. While these findings hold promise, it is necessary to conduct further studies to evaluate the system's performance in vivo and ensure its long-term safety. This research marks a significant stride toward optimizing the efficacy of titanium implants through tailored surface modifications.

Neuromorphic Engineering in Wetware: Discriminating Acoustic Frequencies through Their Effects on Chemical Waves.

Some features of the human nervous system can be mimicked not only through software or hardware but also through liquid solutions of chemical systems maintained under out-of-equilibrium conditions. We describe the possibility of exploiting a thin layer of the Belousov-Zhabotinsky (BZ) reaction as a surrogate for the cochlea for sensing acoustic frequencies. Experiments and simulations demonstrate that, as in the human ear where the cochlea transduces the mechanical energy of the acoustic frequencies into the electrochemical energy of neural action potentials and the basilar membrane originates topographic representations of sounds, our bioinspired chemoacoustic system, based on the BZ reaction, gives rise to spatiotemporal patterns as the representation of distinct acoustic bands through transduction of mechanical energy into chemical energy. Acoustic frequencies in the range 10-2000 Hz are partitioned into seven distinct bands based on three attributes of the emerging spatiotemporal patterns: (1) the types and frequencies of the chemical waves, (2) their velocities, and (3) the Faraday waves' wavelengths.

Proposal of Smith-Waterman algorithm on FPGA to accelerate the forward and backtracking steps.

In bioinformatics, alignment is an essential technique for finding similarities between biological sequences. Usually, the alignment is performed with the Smith-Waterman (SW) algorithm, a well-known sequence alignment technique of high-level precision based on dynamic programming. However, given the massive data volume in biological databases and their continuous exponential increase, high-speed data processing is necessary. Therefore, this work proposes a parallel hardware design for the SW algorithm with a systolic array structure to accelerate the forward and backtracking steps. For this purpose, the architecture calculates and stores the paths in the forward stage for pre-organizing the alignment, which reduces the complexity of the backtracking stage. The backtracking starts from the maximum score position in the matrix and generates the optimal SW sequence alignment path. The architecture was validated on Field-Programmable Gate Array (FPGA), and synthesis analyses have shown that the proposed design reaches up to 79.5 Giga Cell Updates per Second (GCPUS).

Morphogenesis in Synthetic Chemical Cells.

We present an efficient model for describing morphogenesis and the emergence of spatiotemporal structures in synthetic chemical cells. This work is motivated by an experimental setup used for testing Turing's theory of morphogenesis. The model developed is based on the general theory of chemically active droplets, which combines the classical theory of phase separation with reaction-diffusion systems. Through the 2D calculations, we find the six spatiotemporal structures predicted by Turing in 1952 and experimentally observed, in a 1D array of droplets. Moreover, under Turing instability, with a determined chemical wavelength, the system undergoes morphogenesis. This theoretical approach provides a useful tool for understanding the physical differentiation through the direct calculation of the osmotic pressure in each cell as the chemical reaction occurs.

Revisiting the hydroxylation phenomenon of SiO2: a study through "hard-hard" and "soft-soft" interactions.

Surface hydroxylation has been extensively studied over the years for a variety of applications, and studies involving hydroxylation of different silica surfaces are still carried out due to the interesting properties obtained from those modified surfaces. Although a number of theoretical studies have been employed to evaluate details on the hydroxylation phenomenon on silica (SiO2) surfaces, most of these studies are based on computationally expensive models commonly based on extended systems. In order to circumvent such an aspect, here we present a low-cost theoretical study on the SiO2 hydroxylation process aiming to evaluate aspects associated with water-SiO2 interaction. Details about local reactivity, chemical softness, and electrostatic potential were evaluated for SiO2 model substrates in the framework of the density functional theory (DFT) using a molecular approach. The obtained results from this new and promising approach were validated and complemented by fully atomistic reactive molecular dynamics (FARMD) simulations. Furthermore, the implemented approach proves to be a powerful tool that is not restricted to the study of hydroxylation, opening a promising route for low computational cost to analyze passivation and anchoring processes on a variety of oxide surfaces.

Polyamines, IAA and ABA during germination in two recalcitrant seeds: Araucaria angustifolia (Gymnosperm) and Ocotea odorifera (Angiosperm).

BACKGROUND AND AIMS: Plant growth regulators play an important role in seed germination. However, much of the current knowledge about their function during seed germination was obtained using orthodox seeds as model systems, and there is a paucity of information about the role of plant growth regulators during germination of recalcitrant seeds. In the present work, two endangered woody species with recalcitrant seeds, Araucaria angustifolia (Gymnosperm) and Ocotea odorifera (Angiosperm), native to the Atlantic Rain Forest, Brazil, were used to study the mobilization of polyamines (PAs), indole-acetic acid (IAA) and abscisic acid (ABA) during seed germination. METHODS: Data were sampled from embryos of O. odorifera and embryos and megagametophytes of A. angustifolia throughout the germination process. Biochemical analyses were carried out in HPLC. KEY RESULTS: During seed germination, an increase in the (Spd + Spm) : Put ratio was recorded in embryos in both species. An increase in IAA and PA levels was also observed during seed germination in both embryos, while ABA levels showed a decrease in O. odorifera and an increase in A. angustifolia embryos throughout the period studied. CONCLUSIONS: The (Spd + Spm) : Put ratio could be used as a marker for germination completion. The increase in IAA levels, prior to germination, could be associated with variations in PA content. The ABA mobilization observed in the embryos could represent a greater resistance to this hormone in recalcitrant seeds, in comparison to orthodox seeds, opening a new perspective for studies on the effects of this regulator in recalcitrant seeds. The gymnosperm seed, though without a connective tissue between megagametophyte and embryo, seems to be able to maintain communication between the tissues, based on the likely transport of plant growth regulators.

A full-parallel implementation of Self-Organizing Maps on hardware.

Self-Organizing Maps (SOMs) are extensively used for data clustering and dimensionality reduction. However, if applications are to fully benefit from SOM based techniques, high-speed processing is demanding, given that data tends to be both highly dimensional and yet "big". Hence, a fully parallel architecture for the SOM is introduced to optimize the system's data processing time. Unlike most literature approaches, the architecture proposed here does not contain sequential steps - a common limiting factor for processing speed. The architecture was validated on FPGA and evaluated concerning hardware throughput and the use of resources. Comparisons to the state of the art show a speedup of 8.91× over a partially serial implementation, using less than 15% of hardware resources available. Thus, the method proposed here points to a hardware architecture that will not be obsolete quickly.

Of Our Territory, We Know: social mapping experience for emergencies and disasters. / De Nosso Território Sabemos Nós: experiência de cartografia social para emergências e desastres.

The article presents the project "Of Our Territory, We Know" (De Nosso Território Sabemos Nós), carried out in two communities in the mountain cities of the state of Rio de Janeiro that suffered the 2011 disaster, aiming at creating a device for online communication, associated with social cartography, seeking to establish a dialogue between the city management and citizens in a permanent and dynamic way. The device allows local needs to be visible for the creation of public policies. The implementation of the Extended Research Communities (Comunidades Ampliadas de Pesquisa), consisting of residents and their associations, participants of the municipal management, and researchers in a regime of ecology of knowledges, took place through cartographic workshops. The identification of the vulnerabilities and potentials of the communities, the emergence of memories and local knowledges allow the strengthening of community resilience. Therefore, the appraisal of the experience in the creation of social cartographies brings out the group's self-awareness. The mapping process demonstrated how unknown the territories are to the municipal management and even to their inhabitants.

O artigo apresenta o projeto De Nosso Território Sabemos Nós, em duas comunidades das cidades serranas fluminenses que sofreram o desastre de 2011, com o objetivo da criação de dispositivo para comunicação on-line, associado à cartografia social, buscando pôr em diálogo gestão e cidadãos de maneira permanente e dinâmica. O dispositivo permite que as necessidades locais sejam visíveis para deliberação de políticas públicas. A constituição das Comunidades Ampliadas de Pesquisa, composta por moradores e suas associações, participantes da gestão municipal e pesquisadores em regime de ecologia dos saberes, se realizaram através de oficinas cartográficas. A identificação das vulnerabilidades e potencialidades das comunidades, o aflorar das memórias e conhecimentos locais permitem o fortalecimento da resiliência comunitária. Dessa forma, a valorização da experiência na elaboração das cartografias sociais faz emergir a autoconsciência do grupo. O processo de mapeamento demonstrou o quanto os territórios são desconhecidos para a gestão municipal e até mesmo para seus moradores.

New details of assembling bioactive films from dispersions of amphiphilic molecules on titania surfaces.

Tailoring the surface properties of materials for biomedical applications is important to avoid clinical complications. Forming thin layers of amphiphilic molecules with apolar regions that facilitate attractive intermolecular interactions, can be a suitable and versatile approach to achieve hydrophobic surface modification and provide functional antibacterial properties. Aiming to correlate layer structure and properties starting from film formation, octadecylphosphonic acid (ODPA) and dimethyloctadecyl (3-trimethoxysilylpropyl) ammonium chloride (DMOAP) layers were adsorbed onto smooth titania surfaces. Then the films were studied by atomic force microscopy (AFM) and X-ray Photoelectron Spectroscopy (XPS), and their interactions with aqueous environments were characterized by contact angle and zeta potential measurements. In addition, antibacterial assays were performed using E. coli and S. mutants to reveal the antibacterial properties effected by the surface modification. Immediately after sputter deposition, titania was hydrophilic; however, after air storage and adsorption of DMOAP or ODPA, an increase in the water contact angle was observed. XPS investigations after layer formation and after antibacterial tests revealed that the attachment of layers assembled from ODPA on titania substrates is considerably stronger and more stable than that observed for DMOAP films. Heat treatment strongly affects DMOAP layers. Furthermore, DMOAP layers are not stable under biological conditions.

Oncological Safety of Lipofilling in Healthy BRCA Carriers After Bilateral Prophylactic Mastectomy: A Case Series.

OBJECTIVE: The germline breast cancer gene (BRCA) mutation confers a lifetime high risk for breast cancer (BC) and bilateral prophylactic mastectomy is the procedure which allows the highest risk reduction rate. Among other techniques, lipofilling (LF) can be used for breast reconstruction of these patients. However, there are some oncological safety concerns on the subject. The purpose of this study was to assess the oncological risk of LF in BRCA healthy patients. MATERIALS AND METHODS: A single institution case series was built including BRCA I/II mutated patients with no previous history of BC, who underwent bilateral prophylactic mastectomy followed by breast reconstruction with exclusive LF or combined with implants or latissimus dorsi flap. Data were collected regarding patient demographics, clinical information, reconstruction techniques used, and fat grafting details. RESULTS: From September 1999 till November 2017, we identified 18 BRCA carriers with no history of BC who had undergone bilateral prophylactic mastectomy, followed by breast reconstruction with LF. A total of 36 LF procedures were performed following an implant or latissimus dorsi flap, or as an exclusive fat grafting breast reconstruction. The average number of LF sessions was 1.4 with a mean volume of 108.8cc per breast. Median follow-up was 33.0 months after mastectomy and 24.5 months after the last LF intervention; no patients were diagnosed with BC during follow-up. CONCLUSION: Germline BRCA mutation is a high-risk plight for BC. However, despite the limited follow-up, no BC was detected.

De novo assembly and transcriptome of Pfaffia glomerata uncovers the role of photoautotrophy and the P450 family genes in 20-hydroxyecdysone production.

Pfaffia glomerata is a medically important species because it produces the phytoecdysteroid 20-hydroxyecdysone (20-E). However, there has been no ready-to-use transcriptome data available in the literature for this plant. Here, we present de novo transcriptome sequencing of RNA from P. glomerata in order to investigate the 20-E production as well as to understand the biochemical pathway of secondary metabolites in this non-model species. We then analyze the effect of photoautotrophy on the production of 20-E genes phylogenetically identified followed by expression analysis. For this, total messenger RNA (mRNA) from leaves, stems, roots, and flowers was used to construct indexed mRNA libraries. Based on the similarity searches against plant non-redundant protein database, gene ontology, and eukaryotic orthologous groups, 164,439 transcripts were annotated. In addition, the effect of photoautotrophy in two genes putatively involved in the 20-E synthesis pathway was analyzed. The Phantom gene (CYP76C), a precursor of the route, showed increased expression in P. glomerata plants cultured under photoautotrophic conditions. This was accompanied by increased production of this metabolite indicating a putative involvement in 20-E synthesis. This work reveals that several genes in the P. glomerata transcriptome are related to secondary metabolism and stresses, that genes of the P450 family participate in the 20-E biosynthesis route, and that plants cultured under photoautotrophic conditions promote an upregulated Phantom gene and enhance the productivity of 20-E. The data will be used for future investigations of the 20-E synthesis pathway in P. glomerata while offering a better understanding of the metabolism of the species.

Building the National Database of Health Centred on the Individual: Administrative and Epidemiological Record Linkage - Brazil, 2000-2015.

INTRODUCTION: In Brazil, the National Health System (SUS) provides healthcare to the public. The system has multiple administrative databases; the major databases record hospital (SIH) and outpatient (SIA) procedures. Epidemiological information is collected for all populations in subsystems, such as mortality (SIM), live births (SINASC) and diseases of compulsory declaration (SINAN). Each subsystem has its own information system, which is able to provide information about consultations, clinical information and medicines dispensed. However, these systems are not linked, thereby preventing individual-centred analysis. OBJECTIVE: To describe the methods and results of parameter setting that are needed to execute the probabilistic deduplication of large administrative and epidemiological databases in Brazil and to create a National Health Database Centred on the individual. METHODS: This paper shows the results of a record linkage model to integrate data from SIH, SIA, SIM, and SINAN, which have different formats and attributes between them and over time. These data consist of 1.3 billion records from 2000-2015. Probabilistic and deterministic record linkages were used to deduplicate these data. The Kappa statistic and clerical review were used to ensure the quality of the linkage. The graph algorithm and depth-first search were used to generate the identifiers. RESULTS: The deterministic deduplication process resulted in a database with 403,113,527 possible unique individuals. After the probabilistic deduplication process of the former database was performed, 159,703,805 unique individuals were identified. This result had an estimated a false positive error rate of 3.3%, and the false negative error was estimated at 12.3%. CONCLUSIONS: The National Health Database centred on the individual was generated and will allow researchers to use real-world evidence to conduct clinical, epidemiological, economic and other studies. This database represents a significant cohort, spanning 15 years of historical data and preserving patient privacy. The success of the process described will allow repeating and appending the data for future years and enable important studies to promote SUS efficiency and provide better treatments for patients.