This second edition offers a comprehensive overview of the physiological functions of vertebrate kidneys from a comparative viewpoint, with particular emphasis on nonmammalian vertebrates. The topics covered include renal structure; glomerular ultrafiltration; tubular transport of inorganic ions, organic substances, and fluid; and urine dilution and concentration. Mammalian renal function is only considered for purposes of comparison with nonmammalian renal function and as a frame of reference for some of the discussions. The major findings on nonmammalian renal function and the important unanswered questions raised by those findings are described in detail. As such, the book provides comprehensive information on comparative renal function for biological scientists and advanced students of biology with some knowledge of physiology and a desire to know more about renal function in vertebrates, and for mammalian renal physiologists who wish to obtain a broader view of renal function.

Investigation of the mechanisms of cellular response to different mechanical stimuli, as well as mechano-electrical feedback (MEF) in the intact heart is one of the main topics in fundamental and clinical cardiology. The present volume of "Mechanosensitivity in Cells and Tissues: Mechanosensitivity of the Heart" c- bines excellent reviews written by worldwide leaders in this ?eld. The 3rd volume is a great addition to this excellent series of books edited by Andre Kamkin and Irina Kiseleva. This volume successfully combines reviews, aimed at academic, physiology and clinical cardiology communities, devoted to mechanosensitivity of the normal and diseased heart at the ion channel, cell, tissue and organ levels. Kamkin and Kiseleva have made signi?cant contributions to the investigation of mechanosentive ion channels in cardiomyocytes and ?broblasts. Their ba- ground, in addition to extensive collaborations helped them to ?nd and consolidate valuable research ?ndings from prominent specialists in the ?eld of cardiac mechanosensitivity. In the last decade, interest in the role of MEF in the heart has increased sign- cantly. MEF within cardiac tissue is a complex phenomenon in which electroph- iological changes are triggered by myocardial stretch. This phenomenon has been studiedintheclinicalcommunityforoveracenturyandmayhavebothpro-rhythmic and arrhythmogenic consequences. While signi?cant advances have been made in understanding of the effects of mechanical forces on cardiac cells, many questions remain regarding the mechanisms whereby mechanical forces are transduced into changes which alter the behavior of various cardiac cells.

The dynamic field of flavin and flavoprotein biochemistry has seen rapid advancement in recent years. This comprehensive two volume set provides an overview of all aspects of contemporary research in this important class of enzymes. Topics treated include flavoproteins involved in energy generation, signal transduction and electron transfer (including respiration); oxygen activation by flavoproteins; the biology and biochemistry of complex flavoproteins; flavin and flavoprotein photochemistry/photophysics as well as biotechnological applications of flavoproteins. Recent developments in this field include new structures (including those of large membrane-integral electron transfer complexes containing FMN or FAD), elucidation of the role of flavoproteins in cell signalling pathways (including both phototaxis and the circadian cycle) and important new insights into the reaction mechanisms of flavin-containing enzymes. This volume focussing on oxidases, dehydrogenases and related systems is an essential reference for all researchers in biochemistry, chemistry, photochemistry and photophysics working on flavoenzymes.

Following three printings of the First Edition (1978), the publisher has asked for a Second Edition to bring the contents up to date. In doing so the authors aim to show how the newer microscopies are related to the older types with respect to theoretical resolving power (what you pay for) and resolution (what you get). The book is an introduction to students, technicians, technologists, and scientists in biology, medicine, science, and engineering. It should be useful in academic and industrial research, consulting, and forensics; how ever, the book is not intended to be encyclopedic. The authors are greatly indebted to the College of Textiles of North Carolina State University at Raleigh for support from the administration there for typing, word processing, stationery, mailing, drafting diagrams, and general assistance. We personally thank Joann Fish for word process ing, Teresa M. Langley and Grace Parnell for typing services, Mark Bowen for drawing graphs and diagrams, Chuck Gardner for photographic ser vices, Deepak Bhattavahalli for his work with the proofs, and all the other people who have given us their assistance. The authors wish to acknowledge the many valuable suggestions given by Eugene G. Rochow and the significant editorial contributions made by Elizabeth Cook Rochow."

Since the initial establishment of Robert Koch's postulates in the nineteenth century, microbial protein toxins have been recognized as a major factor of bacterial and fungal virulence. An increasing number of proteins produced and secreted by various bacteria, yeasts and plants are extremely toxic and most of them developed remarkably "intelligent" strategies to enter, to penetrate and to finally kill a eukaryotic target cell by modifying or blocking essential cellular components.

This book describes the strategies employed by protein toxins to render their pro- and eukaryotic producers a selective growth advantage over competitors. In providing an up-to-date overview on the mode of protein toxin actions, it accommodates biomedically and biologically relevant toxin model systems. As a result, it significantly broadens our perspective on biochemical architecture and molecular ploy behind the lethal principles of pro- and eukaryotic toxins.

Ligand-gated ion channels mediate fast synaptic transmission in the central nervous s- tem (CNS) and at ganglionic and neuromuscular synapses. The nicotinic acetylcholine - ceptor (nAChR) is a member of the ligand-gated ion channel superfamily, which includes the 5-HT , glycine and GABA type A and C receptors. These receptors are known as Cys- 3 loop receptors, as all of them have a conserved sequence containing a pair of cysteines separated by 13 residues and linked by a disulfide bridge. nAChRs can be divided into two groups: muscle receptors, which are found at the skeletal neuromuscular junction where they mediate neuromuscular transmission, and neuronal receptors, which are found throughout the peripheral and central nervous system. Many of the early studies carried out on the subunit composition and structure of the nAChRs were performed on receptors isolated from the electric organ of Torpedo californica, as this tissue is very rich in nAChRs, and they were found to have a high degree of homology with the embryonic v- tebrate muscle type receptor. Muscle nAChRs are made up of five subunits arranged around a central pore (Fig. 1A, B). In Torpedo electric organ and vertebrate fetal muscle, the subunit composition is (a1) b1gd, and in adult muscle the g subunit is replaced by the e to give an (a1)b1ed 2 2 composition (Mishina et al. 1986).

Time-resolved fluorescence spectroscopy is widely used as a research tool in bioch- istry and biophysics. These uses of fluorescence have resulted in extensive knowledge of the structure and dynamics of biological macromolecules. This information has been gained by studies of phenomena that affect the excited state, such as the local environment, quenching processes, and energy transfer. Topics in Fluorescence Spectroscopy, Volume 4: Probe Design and Chemical Sensing reflects a new trend, which is the use of time-resolved fluorescence in analytical and clinical chemistry. These emerging applications of time-resolved fluorescence are the result of continued advances in laser detector and computer technology. For instance, pho- multiplier tubes (PMT) were previously bulky devices. Miniature PMTs are now available, and the performance of simpler detectors is continually improving. There is also considerable effort to develop fluorophores that can be excited with the red/ne- infrared (NIR) output of laser diodes. Using such probes, one can readily imagine small time-resolved fluorometers, even hand-held devices, being used fordoctor's office or home health care.

Phytochemicals from medicinal plants are receiving ever greater attention in the scientific literature, in medicine, and in the world economy in general. For example, the global value of plant-derived pharmaceuticals will reach $500 billion in the year 2000 in the OECD countries. In the developing countries, over-the-counter remedies and "ethical phytomedicines," which are standardized toxicologically and clinically defined crude drugs, are seen as a promising low cost alternatives in primary health care. The field also has benefited greatly in recent years from the interaction of the study of traditional ethnobotanical knowledge and the application of modem phytochemical analysis and biological activity studies to medicinal plants. The papers on this topic assembled in the present volume were presented at the annual meeting of the Phytochemical Society of North America, held in Mexico City, August 15-19, 1994. This meeting location was chosen at the time of entry of Mexico into the North American Free Trade Agreement as another way to celebrate the closer ties between Mexico, the United States, and Canada. The meeting site was the historic Calinda Geneve Hotel in Mexico City, a most appropriate site to host a group of phytochemists, since it was the address of Russel Marker. Marker lived at the hotel, and his famous papers on steroidal saponins from Dioscorea composita, which launched the birth control pill, bear the address of the hotel."

Advances in biochemistry, cell biology, genome-wide mutagenesis - coupled with molecular technology, including gene microarray and transgenic and knock-out animals - have been instrumental in understanding the cellular processes and molecular pathways of self-tolerance and autoimmune diseases. The molecular definition of these pathways and processes has led to novel treatments for certain auto-immune diseases that are based on the pathogenesis of diseases rather than on broad-spectrum immunosuppression. This book reviews many of these current developments and proposes future novel approaches for understanding the pathogenesis of auto-immune diseases and designing novel therapy. This book covers three major areas of auto-immunity: the basic mechanisms of immunological tolerance, pathogenesis of auto-immune diseases, and some novel therapies. This book should be useful for immunologists, molecular biologists, rheumatologists, and clinical scientists.

''Interesting with many useful ideas and references. It covers a broad range and it is a good introduction to this field.'' ---Analyst

Transposable elements are short lengths of DNA with the capacity to move between different points within a genome. This process can affect the function of genes at or near the insertion site. The present book gives an overview of the impact of transposable elements on plant genomes and explains how to recognize and study transposable elements, e.g. by using state-of-the-art strategies like "new generation sequencing." Moreover, the impact of transposable elements on plant genome structure and function is reviewed in detail, and also illustrated in examples and case studies. The book is intended both for readers familiar with the field and for newcomers. With large-scale sequencing becoming increasingly available, more and more people will come across transposable element sequences in their data, and this volume will hopefully help to convince them that they are not just "junk DNA."

The present book collects selected contributions from researchers working in the field of food science, and participating at the second spring school for Food Quality, Safety and Technology, which was held in Botucatu (Sao Paulo, Brazil), from September 24th to 27th, 2012, at the Botucatu Campus of the Universidade Estadual Paulista Julio Mesquita Filho (UNESP). The goal of the conference was to provide a scientific forum covering large areas of agronomy, nutrition, food science and technology, veterinary and other areas related to food technology development. Teachers, professionals, graduate and post-graduate students in Food Science; Food and Agriculture Engineering; Veterinary, Science and Food Technology and related areas were addressed by providing an exchange of knowledge and technologies. The initiative aimed to establish uniform, globally recognized scientific principles on food safety and quality, which could be consistently applied to industry and production sectors and stakeholders, taking into account that effective food control systems are essential to protecting the health and safety of domestic consumers, to guaranteeing the safety and quality of foods entering international trade, and to ensuring that imported foods conform to national requirements."

This volume focuses and describes tools, assays, and techniques used to enhance the understanding of the role of auxins and cytokinins. The chapters in this book cover topics such as: microbial manipulation of auxin and cytokinins in plants; interplay between auxin and cytokinin and its impact on MAPK; H2O2 production in Arabidopsis leaves; crosstalk between jasmonate and auxin in plant stress responses of roots; and high-throughput protoplast trans-activation (PTA) screening. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Cutting-edge and comprehensive, Auxins and Cytokinins in Plant Biology: Methods and Protocols is a valuable resource to plant scientists, graduate and under graduate students in addressing their biological questions relevant to the functional implications of auxin and cytokinins.

The International Symposium on Prostaglandins and Related Compounds, first held in Vienna 1972, revisited the city after 24 years for the 10TH Symposium. For the many re searchers working in this multi-disciplinary field it was an opportunity to exchange their ex periences and share new data with colleagues from all around the world. This scientific exchange was largely encouraged by the unseasonably cold and rainy weather. For the first time, there was quite a large attendance from the former Communist countries. Eugene Garfield prepared a key note address delivered during the meeting (The Sci entist 1996, 12) reviewing the contribution of the Nobel Laureates U.S. von Euler, l.R. Vane, S.K. Bergstrom, and B.I. Samuelsson, discussing the relevance of the more than 40,000 pa pers in this area published since 1991. Overall, there is still a rapidly growing interest, and in particular a great variety of clinical applications of this family of compounds which were dis cussed in detail during the meeting. Beside the lectures there were 19 workshops covering nearly all the topics of key in terest. All the speakers were invited to prepare a manuscript which has resulted in the volume now in your hands. Special thanks to Dr. Patrick Wong and the new publisher of this series who helped to publish the proceedings in the usual quality and reasonable period of time. Looking forward to seeing all of you again in Florence in 1999, hopefully with much more sun."

Representing the latest knowledge of the ecology and the physiology of cold-adapted microorganisms, plants and animals, this book explains the mechanisms of cold-adaptation on the enzymatic and molecular level, including results from the first crystal structures of enzymes of cold-adapted organisms.

There is a growing need in both industrial and academic research to obtain accurate quantitative results from continuous wave (CW) electron paramagnetic resonance (EPR) experiments. This book describes various sample-related, instrument-related and software-related aspects of obtaining quantitative results from EPR expe- ments. Some speci?c items to be discussed include: selection of a reference standard, resonator considerations (Q, B, B ), power saturation, sample position- 1 m ing, and ?nally, the blending of all the factors together to provide a calculation model for obtaining an accurate spin concentration of a sample. This book might, at ?rst glance, appear to be a step back from some of the more advanced pulsed methods discussed in recent EPR texts, but actually quantitative "routine CW EPR" is a challenging technique, and requires a thorough understa- ing of the spectrometer and the spin system. Quantitation of CW EPR can be subdivided into two main categories: (1) intensity and (2) magnetic ?eld/mic- wave frequency measurement. Intensity is important for spin counting. Both re- tive intensity quantitation of EPR samples and their absolute spin concentration of samples are often of interest. This information is important for kinetics, mechanism elucidation, and commercial applications where EPR serves as a detection system for free radicals produced in an industrial process. It is also important for the study of magnetic properties. Magnetic ?eld/microwave frequency is important for g and nuclear hyper?ne coupling measurements that re?ect the electronic structure of the radicals or metal ions.

Plant-Microbe Interactions, Volume 1 Many plant-microbe interactions have agronomic importance because of either beneficial (e.g., nitrogen fixation or biocontrol) or detrimental (e.g., pathogen esis) effects. Although these systems have been the subjects of scientific re search for many years, recently there has been a tremendous increase in our knowledge of them. The increases in this research have followed a similar general increase in plant science research. Classical plant science research disciplines (e.g., agronomy, breeding, plant physiology, systematics, etc.) have been affected by an increased focus on molecular biology. These new technologies, as well as advances in other areas, have the effect of blurring the traditional borders between research disciplines. Another factor influencing the development of this research is the increased attention given to environmental issues. These concerns have been brought about by debate over the release of genetically modified organisms and the general concern over environmental quality. Thus, research areas focused on plant-microbe interactions are presently in a period of great excitement and growth that shows every sign of continuing far into the future. As in most research areas, the rate of advance and breadth of disciplines involved in the study of plant-microbe interactions make it impossible for the average researcher or student to stay abreast of the primary scientific literature."

Modeling of Oxygen Transport to Skeletal Muscle: Blood Flow Distribution, Shunt, and Diffusion.- The Haldane Effect of Rabbit Blood Under Different Acid-Base Conditions.- Interaction of Blood Flow, Diffusive Transport and Cell Metabolism in Isovolemic Anemia.- The Role of Wall Shear Stress in Microvascular Network Adaptation.- Changes in Tissue Histology Associated with Adaptation and Acclimation to Hypoxia.- Transcutaneous Measurements of Skin O2 Supply and Blood Gases.- Relating Measuring Signals from PO2 Electrodes to Tissue PO2: A Theoretical Study.- A New Catheter for Quasi-Continuous Measurement of Arterial Partial Oxygen Pressure.- Continuous Intra-Arterial PO2 Monitoring During Thoracic Surgery.- Construction, Calibration and Evaluation of PO2 Electrodes for Chronical Implantation in the Rabbit Brain Cortex.- Photothrombosis in Rabbit Brain Cortex: Follow Up by Continuous pO2 Measurement.- Intravitreal and Intraretinal Oxygen Tension in the Rat Eye.- Brain Oxygenation State: Preparation of Isolated Perfused Rat Brain and Near-Infrared Spectrophotometry.- The Simultaneous Measurement of the Redox State of Cytochrome Oxidase in Heart and Brain of Rat In Vivo by NIR.- Picosecond Time of Flight Measurement of Living Tissue: Time Resolved Beer-Lambert Law.- Cerebral Oxygenation State in Chemically-Induced Seizures in the Rat: Study by Near Infrared Spectrophotometry.- Experimentally Measured Optical Pathlengths for the Adult Head, Calf and Forearm and the Head of the Newborn Infant as a Function of Inter Optode Spacing.- Near-Infrared Imaging in vivo (I): Image Restoration Technique Applicable to the NIR Projection Images.- Muscle Oxygenation by Fast Near Infrared Spectrophotometry (NIRS) in Ischemic Forearm.- Near-Infrared Imaging in vivo (II): 2-Dimensional Visualization of Tissue Oxygenation State.- Monitoring of the Oxygen Pressure in the Blood of Live Animals Using the Oxygen Dependent Quenching of Phosphorescence.- Nuclear Magnetic Resonance Spectroscopy and the Study of Tissue Oxygen Metabolism: A Review.- On-Line Oxygen Uptake Measurement (VO2): A Computer Feed-Back Controlled Rebreathing Circuit for Long Term Oxygen Uptake Registration.- Oxygen Delivery (DEL O2) Dependent and Independent Oxygen Consumption (VO2).- Microtopographic Analysis of Oxidative Stress in Organ Microcirculatory Units.- Cytochrome P-450 Under Conditions of Oxidative Stress: Role of Antioxidant Recycling in the Protection Mechanisms.- Ischemia-Reperfusion Injury and Free Radical Involvement in Gastric Mucosal Disorders.- Influence of Anti-Inflammatory Drugs and Free Radical Scavengers on Intestinal Ischemia Induced Oxidative Tissue Damage.- Influence of Free Radical Scavengers on Myeloperoxidase Activity and Lipid Peroxidation in Acute Skin Grafts.- Morphometric Methods for the Evaluation of Capillary Grouping Patterns in Rat Heart.- Estimation of the Oxygen Gradient Across Phospholipid Bilayers of Mitochondria from Reperfused Rabbit Hearts after Ischemia.- The Oxygen Dependence of Mitochondrial Oxidative Phosphorylation and Its Role in Regulation of Coronary Blood Flow.- Oxygen Penetration in and Release from Lung Surfactant.- Improvement of Pulmonary Gas Exchange After Surfactant Replacement in Rats With Pneumocystis Carinii Pneumonia.- Attenuation of Hypoxic Pulmonary Vasoconstriction in Acute Oleic Acid Lung Injury: Significance of Vasodilator Prostanoids.- 99mTc-DTPA Clearance: A Sensitive Method for Early Detection of An Impending Disturbance in Gas Exchange.- Acute Respiratory Failure During Pneumonia Induced by Sendai Virus.- Estimation of Respiratory Mechanics in Dogs with Acute Lung Injury.- Effect of Hyperventilation on Oxygenation of the Brain Cortex of Neonates.- Hypoxia and the "Reaction Theory" of Central Respiratory Chemosensitivity.- Oxygenation of Mammary Tumors: From Isotransplanted Rodent Tumors to Primary Malignancies in Patients.- Measurements of Tumor Blood Flow Using Intraperitoneal Deuterium and 2H-NMR Spectroscopy....

Antimicrobial peptides have been the subject of intense research in the past decades, and are now considered as an essential part of the defense system in bacteria, plants, animals and humans. This book provides an update on these effector molecules of the innate immune system both for researchers who are already actively involved in the area, and for those with a general interest in the topic. The book starts with an overview of the evolution of cysteine- containing antimicrobial peptides (including defensins), and the role of these peptides in host defense in plants and micro- organisms. The realization that antimicrobial peptides also display functions distinct from their direct antimicrobial action is the focus of the next chapters, and puts these peptides center stage in immunity and wound repair. Further chapters discuss the role of antimicrobial peptides in disease, by providing an overview of mechanisms in bacterial resistance to antimicrobial peptides and a discussion of their role in inflammatory bowel disease, cystic fibrosis lung disease and chronic obstructive pulmonary disease. Finally, the book shows how knowledge of the function of antimicrobial peptides and their regulation can be used to design new therapies for inflammatory and infectious disorders. This is a very important area of research because of the increase in resistance of micro-organisms to conventional antibiotics. Therefore the use of synthetic or recombinant peptides, or agents that stimulate the endogenous production of antimicrobial peptides, provides an attractive alternative for conventional antibiotics.

Chemistry for Sustainable Development in Africa gives an insight into current Chemical research in Africa. It is edited and written by distinguished African scientists and includes contributions from Chemists from Northern, Southern, Western, Eastern, Central and Island state African Countries. The core themes embrace the most pressing issues of our time, including Environmental Chemistry, Renewable Energies, Health and Human Well-Being, Food and Nutrition, and Bioprospecting and Commercial Development. This book is invaluable for teaching and research institutes in Africa and worldwide, private sector entities dealing with natural products from Africa, as well as policy and decision-making bodies and non-governmental organizations.

The proceedings of the NATO Advanced Research Workshop on Biological Electron Transfer Chains contains reports on three themes. The first theme concerns electron transfer. The analysis of ET at the molecular level is still fundamental to an understanding of how ET chains operate "in vivo". The second theme concerns bacterial redox chains. These contributions show how complicated these chains can be, often involving a number of gene clusters, each of which in turn consists of an extensive array of genes. The third theme relates to the structure and function of redox proteins. Structural information on proteins is necessary to a proper understanding of their function and of the context in which they operate. The two final chapters deal with oxido-reductases and the cytochrome oxidase family.

This two volume set introduces the up-to-date high-tech applications of Aggregation-Induced Emission (AIE) luminogens mainly in the areas of biosensing, bioimaging, and biomedicine. The 1st volume covers the applications of AIE materials in biosensing and bioimaging, including the technological utilizations in ionic/biomolecular sensing, bacterial imaging, cell imaging, intracellular microenvironment analysis, advanced optical imaging and multimodality, etc. It is an essential reference for materials scientists, chemists, physicists and biological chemists.

Part I The Nano-Scale Biological Systems in Nature; Molecular bio-motors in living cells - by T. Nishizaka; The form designed by viral genome - by K. Onodera; Part II Detection and Characterization Technology; Atomic force microscopy applied to nano-mechanics of the cell - by A. Ikai; Design, synthesis and biological application of fluorescent sensor molecules for cellular imaging - by K. Kikuchi; Dynamic visualization of cellular signaling - by Q. Ni and J. Zhang; Part III Fabrication Technology; Surface acoustic wave atomizer and electrostatic deposition - by Y. Yamagata; Electrospray deposition of biomolecules by V.N. Morozov; Part IV Processing Technology; Droplet handling - by T.Torii; Integrated microfluidic systems - by S. Kaneda and T. Fujii; Part V Applications; A novel non-viral gene delivery system: Multifunctional envelope-type nano device - by H. Hatakeyama, H. Akita, K. Kogure, and H. Harashima; Biosensors - by M. Saito, H.M. Hiep, N. Nagatani, and E.Tamiya; Micro bioreactors - by Sato and T. Kitamori

This book discusses recent advances in theoretical-computational studies on the biosynthesis of melanin pigment (melanogenesis). These advances are being driven by the development of high-performance computers, new experimental findings, and extensive work on medical applications involving the control of pigmentation and the treatment of challenging dermatological diseases. Understanding the elementary processes involved in chemical reactions at the atomic scale is important in biochemical reaction design for effective control of the pigmentary system. Accordingly, the book focuses on the elementary steps involved in melanogenesis, which crucially affect the composition of the resulting melanin pigment by means of competitive reactions. The book also addresses reactions analogous to melanogenesis, with a focus on o-quinone reactions, which are especially important for understanding melanogenesis-associated cytotoxicity.

This book summarizes the advances made in the last decade in the biophysics, biochemistry, and molecular biology of the enzyme known as Photosystem I, the light-induced plastocyanin: ferredoxin oxidoreductase. Photosystem I participates along with Photosystem II in harvesting solar energy to supply photosynthetic organisms with stored chemical energy in the form of ATP and stored reducing power in the form of NADPH for processes such as metabolism, growth, and reproduction.

This volume is a unique compilation of chapters that include information on molecular architecture, protein-pigment interactions, excitation and electron transfer dynamics, protein-cofactor interactions, kinetics of electron transfer and bioassembly of proteins and cofactors.

The volume begins with a series of historical perspectives that provide a solid background to the field, and ends with information on modelling of light-harvesting and electron transfer reactions, and the evolution of the reaction center. Particular attention is paid to spectroscopy, including the theory of the measurement and the interpretation of the data. The book is intended to be a comprehensive and up-to-date source of background information on the Photosystem I reaction center for seasoned researchers, those who are just entering the field, Ph.D. students, researchers and undergraduates in the fields of biophysics, biochemistry, microbiology, agriculture, and ecology.