The structural analysis of the periplasmic domain of Sinorhizobium meliloti chemoreceptor McpZ reveals a novel fold and suggests a complex mechanism of transmembrane signaling.

Chemotaxis is a fundamental process whereby bacteria seek out nutrient sources and avoid harmful chemicals. For the symbiotic soil bacterium Sinorhizobium meliloti, the chemotaxis system also plays an essential role in the interaction with its legume host. The chemotactic signaling cascade is initiated through interactions of an attractant or repellent compound with chemoreceptors or methyl-accepting chemotaxis proteins (MCPs). S. meliloti possesses eight chemoreceptors to mediate chemotaxis. Six of these receptors are transmembrane proteins with periplasmic ligand-binding domains (LBDs). The specific functions of McpW and McpZ are still unknown. Here, we report the crystal structure of the periplasmic domain of McpZ (McpZPD) at 2.7 Å resolution. McpZPD assumes a novel fold consisting of three concatenated four-helix bundle modules. Through phylogenetic analyses, we discovered that this helical tri-modular domain fold arose within the Rhizobiaceae family and is still evolving rapidly. The structure, offering a rare view of a ligand-free dimeric MCP-LBD, reveals a novel dimerization interface. Molecular dynamics calculations suggest ligand binding will induce conformational changes that result in large horizontal helix movements within the membrane-proximal domains of the McpZPD dimer that are accompanied by a 5 Å vertical shift of the terminal helix toward the inner cell membrane. These results suggest a mechanism of transmembrane signaling for this family of MCPs that entails both piston-type and scissoring movements. The predicted movements terminate in a conformation that closely mirrors those observed in related ligand-bound MCP-LBDs.

Complete Genome Sequence of Curtobacterium sp. Isolated from Surface-Sterilized Germinating Alfalfa Seeds.

We reported the complete genome sequence of a member of the pathogenic Curtobacterium genus. The sample includes a circular 3,955-kb chromosome, a 164-kb megaplasmid and a 42-kb plasmid. This strain was isolated from surface-sterilized alfalfa seeds.

McpT, a Broad-Range Carboxylate Chemoreceptor in Sinorhizobium meliloti.

Chemoreceptors enable the legume symbiont Sinorhizobium meliloti to detect and respond to specific chemicals released from their host plant alfalfa, which allows the establishment of a nitrogen-fixing symbiosis. The periplasmic region (PR) of transmembrane chemoreceptors act as the sensory input module for chemotaxis systems via binding of specific ligands, either directly or indirectly. S. meliloti has six transmembrane and two cytosolic chemoreceptors. However, the function of only three of the transmembrane receptors have been characterized so far, with McpU, McpV, and McpX serving as general amino acid, short-chain carboxylate, and quaternary ammonium compound sensors, respectively. In the present study, we analyzed the S. meliloti chemoreceptor McpT. High-throughput differential scanning fluorimetry assays, using Biolog phenotype microarray plates, identified 15 potential ligands for McpTPR, with the majority classified as mono-, di-, and tricarboxylates. S. meliloti exhibited positive chemotaxis toward seven selected carboxylates, namely, α-ketobutyrate, citrate, glyoxylate, malate, malonate, oxalate, and succinate. These carboxylates were detected in seed exudates of the alfalfa host. Deletion of mcpT resulted in a significant decrease of chemotaxis to all carboxylates except for citrate. Isothermal titration calorimetry revealed that McpTPR bound preferentially to the monocarboxylate glyoxylate and with lower affinity to the dicarboxylates malate, malonate, and oxalate. However, no direct binding was detected for the remaining three carboxylates that elicited an McpT-dependent chemotaxis response. Taken together, these results demonstrate that McpT is a broad-range carboxylate chemoreceptor that mediates chemotactic response via direct ligand binding and an indirect mechanism that needs to be identified. IMPORTANCE Nitrate pollution is one of the most widespread and challenging environmental problems that is mainly caused by the agricultural overapplication of nitrogen fertilizers. Biological nitrogen fixation by the endosymbiont Sinorhizobium meliloti enhances the growth of its host Medicago sativa (alfalfa), which also efficiently supplies the soil with nitrogen. Establishment of the S. meliloti-alfalfa symbiosis relies on the early exchange and recognition of chemical signals. The present study contributes to the disclosure of this complex molecular dialogue by investigating the underlying mechanisms of carboxylate sensing in S. meliloti. Understanding individual steps that govern the S. meliloti-alfalfa molecular cross talk helps in the development of efficient, commercial bacterial inoculants that promote the growth of alfalfa, which is the most cultivated forage legume in the world, and improves soil fertility.

Rhizobial Chemoattractants, the Taste and Preferences of Legume Symbionts.

The development of host-microbe interactions between legumes and their cognate rhizobia requires localization of the bacteria to productive sites of initiation on the plant roots. This end is achieved by the motility apparatus that propels the bacterium and the chemotaxis system that guides it. Motility and chemotaxis aid rhizobia in their competitiveness for space, resources, and nodulation opportunities. Here, we examine studies on chemotaxis of three major model rhizobia, namely Sinorhizobium meliloti, Rhizobium leguminosarum, and Bradyrhizobium japonicum, cataloging their range of attractant molecules and correlating this in the context of root and seed exudate compositions. Current research areas will be summarized, gaps in knowledge discussed, and future directions described.

Using Circular Dichroism to Control Energy Transfer in Multiphoton Ionization.

Chirality causes symmetry breaks in a large variety of natural phenomena ranging from particle physics to biochemistry. We investigate one of the simplest conceivable chiral systems, a laser-excited, oriented, effective one-electron Li target. Prepared in a polarized p state with |m|=1 in an optical trap, the atoms are exposed to co- and counterrotating circularly polarized femtosecond laser pulses. For a field frequency near the excitation energy of the oriented initial state, a strong circular dichroism is observed and the photoelectron energies are significantly affected by the helicity-dependent Autler-Townes splitting. Besides its fundamental relevance, this system is suited to create spin-polarized electron pulses with a reversible switch on a femtosecond timescale at an energy resolution of a few meV.

An Updated Perspective on Sinorhizobium meliloti Chemotaxis to Alfalfa Flavonoids.

The symbiotic interaction between leguminous plants and their cognate rhizobia allows for the fixation of gaseous dinitrogen into bioavailable ammonia. The perception of host-derived flavonoids is a key initial step for the signaling events that must occur preceding the formation of the nitrogen-fixing organ. Past work investigating chemotaxis - the directed movement of bacteria through chemical gradients - of Bradyrhizobium japonicum, Rhizobium leguminosarum, and Rhizobium meliloti discovered chemotaxis to various organic compounds, but focused on chemotaxis to flavonoids because of their relevance to the symbiosis biochemistry. The current work sought to replicate and further examine Sinorhizobium (Ensifer) meliloti chemotaxis to the flavonoids previously thought to act as the principal attractant molecules prior to the initial signaling stage. Exudate from germinating alfalfa seedlings was analyzed for composition and quantities of different flavonoid compounds using mass spectrometry. The abundance of four prevalent flavonoids in germinating alfalfa seed exudates (SEs) was at a ratio of 200:5:5:1 for hyperoside, luteolin, luteolin-7-glucoside, and chrysoeriol. Using quantitative chemotaxis capillary assays, we did not detect chemotaxis of motile S. meliloti cells to these, and two other flavonoids identified in seed exudates. In support of these findings, the flavonoid fraction of seed exudates was found to be an insignificant attractant relative to the more hydrophilic fraction. Additionally, we observed that cosolvents commonly used to dissolve flavonoids confound the results. We propose that the role flavonoids play in S. meliloti chemotaxis is insignificant relative to other components released by alfalfa seeds.

Sinorhizobium meliloti Chemoreceptor McpV Senses Short-Chain Carboxylates via Direct Binding.

Sinorhizobium meliloti is a soil-dwelling endosymbiont of alfalfa that has eight chemoreceptors to sense environmental stimuli during its free-living state. The functions of two receptors have been characterized, with McpU and McpX serving as general amino acid and quaternary ammonium compound sensors, respectively. Both receptors use a dual Cache (calcium channels and chemotaxis receptors) domain for ligand binding. We identified that the ligand-binding periplasmic region (PR) of McpV contains a single Cache domain. Homology modeling revealed that McpVPR is structurally similar to a sensor domain of a chemoreceptor with unknown function from Anaeromyxobacter dehalogenans, which crystallized with acetate in its binding pocket. We therefore assayed McpV for carboxylate binding and S. meliloti for carboxylate sensing. Differential scanning fluorimetry identified 10 potential ligands for McpVPR Nine of these are monocarboxylates with chain lengths between two and four carbons. We selected seven compounds for capillary assay analysis, which established positive chemotaxis of the S. meliloti wild type, with concentrations of peak attraction at 1 mM for acetate, propionate, pyruvate, and glycolate, and at 100 mM for formate and acetoacetate. Deletion of mcpV or mutation of residues essential for ligand coordination abolished positive chemotaxis to carboxylates. Using microcalorimetry, we determined that dissociation constants of the seven ligands with McpVPR were in the micromolar range. An McpVPR variant with a mutation in the ligand coordination site displayed no binding to isobutyrate or propionate. Of all the carboxylates tested as attractants, only glycolate was detected in alfalfa seed exudates. This work examines the relevance of carboxylates and their sensor to the rhizobium-legume interaction.IMPORTANCE Legumes share a unique association with certain soil-dwelling bacteria known broadly as rhizobia. Through concerted interorganismal communication, a legume allows intracellular infection by its cognate rhizobial species. The plant then forms an organ, the root nodule, dedicated to housing and supplying fixed carbon and nutrients to the bacteria. In return, the engulfed rhizobia, differentiated into bacteroids, fix atmospheric N2 into ammonium for the plant host. This interplay is of great benefit to the cultivation of legumes, such as alfalfa and soybeans, and is initiated by chemotaxis to the host plant. This study on carboxylate chemotaxis contributes to the understanding of rhizobial survival and competition in the rhizosphere and aids the development of commercial inoculants.

Sinorhizobium meliloti Chemotaxis to Multiple Amino Acids Is Mediated by the Chemoreceptor McpU.

The legume symbiont Sinorhizobium meliloti is chemoattracted to compounds exuded by germinating seeds of its host alfalfa. This response is mainly mediated by the S. meliloti chemoreceptor McpU. McpU also has a prominent contribution in sensing a synthetic amino acid (aa) mixture mimicking the amounts and composition observed in seed exudate. Here, we used the hydrogel capillary assay to quantify chemotactic responses of S. meliloti to individual aa exuded by germinating alfalfa seeds and to define the role of McpU in this behavior. S. meliloti exhibited positive chemotaxis responses to all proteinogenic aa, except for aspartate, and to citrulline, cystine, gamma-aminobutyric acid, and ornithine. Wild-type responses were diverse in intensity, while a strain lacking mcpU displayed strongly diminished responses. Differential scanning fluorimetry demonstrated interaction of the purified periplasmic region of McpU (McpU-PR) with the aa, except glutamate and aspartate. We additionally tested organic acids and sugars, but there were no significant interactions with the McpU ligand-binding domain, except for citrate. Using ligand displacement, we confirmed the interaction of McpU-PR with aa representing strong and weak attractants. Our results show that S. meliloti McpU is a broad-range aa receptor mediating differential responses to individual attractants, which does not bind negatively charged aa.

Sinorhizobium meliloti chemotaxis to quaternary ammonium compounds is mediated by the chemoreceptor McpX.

The bacterium Sinorhizobium meliloti is attracted to seed exudates of its host plant alfalfa (Medicago sativa). Since quaternary ammonium compounds (QACs) are exuded by germinating seeds, we assayed chemotaxis of S. meliloti towards betonicine, choline, glycine betaine, stachydrine and trigonelline. The wild type displayed a positive response to all QACs. Using LC-MS, we determined that each germinating alfalfa seed exuded QACs in the nanogram range. Compared to the closely related nonhost species, spotted medic (Medicago arabica), unique profiles were released. Further assessments of single chemoreceptor deletion strains revealed that an mcpX deletion strain displayed little to no response to these compounds. Differential scanning fluorimetry showed interaction of the isolated periplasmic region of McpX (McpXPR and McpX34-306 ) with QACs. Isothermal titration calorimetry experiments revealed tight binding to McpXPR with dissociation constants (Kd ) in the nanomolar range for choline and glycine betaine, micromolar Kd for stachydrine and trigonelline and a Kd in the millimolar range for betonicine. Our discovery of S. meliloti chemotaxis to plant-derived QACs adds another role to this group of compounds, which are known to serve as nutrient sources, osmoprotectants and cell-to-cell signalling molecules. This is the first report of a chemoreceptor that mediates QACs taxis through direct binding.

Assessment of potential atmospheric transport and deposition patterns due to Russian Pacific Fleet operations.

A probabilistic analysis of atmospheric transport and deposition patterns from two nuclear risk sites-Kamchatka and Vladivostok-situated in the Russian Far East to countries and geographical regions of interest (Japan, China, North and South Koreas, territories of the Russian Far East, State of Alaska, and Aleutian Chain Islands, US) was performed. The main questions addressed were the following: Which geographical territories are at the highest risk from hypothetical releases at these sites? What are the probabilities for radionuclide atmospheric transport and deposition on different neighboring countries in case of accidents at the sites? For analysis, several research tools developed within the Arctic Risk Project were applied: (1) isentropic trajectory model to calculate a multiyear dataset of 5-day forward trajectories that originated over the site locations at various altitudes; (2) DERMA long-range transport model to simulate 5-day atmospheric transport, dispersion, and deposition of 137Cs for 1-day release (at the rate of 10(10) Bq/s); and (3) a set of statistical methods (including exploratory, cluster, and probability fields analyses) for evaluation of trajectory and dispersion modeling results. The possible impact (on annual, seasonal, and monthly basis) of selected risk sites on neighboring geographical regions is evaluated using a set of various indicators. For trajectory modeling, the indicators examined are: (1) atmospheric transport pathways, (2) airflow probability fields, (3) fast transport probability fields, (4) maximum possible impact zone, (5) maximum reaching distance, and (6) typical transport time fields. For dispersion modeling, the indicators examined are: (1) time integrated air concentration, (2) dry deposition, and (3) wet deposition. It was found for both sites that within the boundary layer the westerly flows are dominant throughout the year (more than 60% of the time), increasing with altitude of free troposphere up to 85% of the time. For the Kamchatka site, the US regions are at the highest risk with the average times of atmospheric transport ranging from 3 to 5.1 days and depositions of 10(-1) Bq/m2 and lower. For the Vladivostok site, the northern China and Japan regions are at the highest risk with the average times of atmospheric transport of 0.5 and 1.6 days, respectively, and depositions ranging from 10(0) to 10(+2) Bq/m2. The areas of maximum potentially impacted zones are 30 x 10(4) km2 and 25 x 10(4) km2 for the Kamchatka and Vladivostok sites, respectively.

Factors affecting cathode ray tube display performance.

Monitors are a complex combination of analog circuits that perform absolutely amazing feats of current and voltage control at both low and high frequency. Digital circuits can reduce the complexity of interfacing with the adjustments and even be used to build complex waveforms, but they cannot replace the point at which pure power is required. Five main topic areas are covered, each building on the other in how they relate to display performance. They are glass composition, resolution and addressability, phosphor selection and aging characteristics, light output and uniformity, and the tools available to assess display performance.

Individual interventions to prevent drunk driving: types, efficacy, and a theoretical perspective.

College students (n = 100) who had tried to stop someone from driving drunk, or who someone else had tried to stop, provided information about their interaction, including what was said and whether the intervention worked. Results suggest that the manner in which people intervene can affect the likelihood that the impaired person will not drive, with forceful statements, clear demands, and concrete action being more effective than requests, pleas, or suggestions. The hypothesis that intervention represents a threat to the person's image received limited support, possibly for methodological reasons. The concept of threat to competence is discussed, as are implications of the results for prevention and education activities.

Induction of glutathione s-transferase-pi in Barrett's metaplasia and Barrett's adenocarcinoma cell lines.

Barrett's metaplasia consists of columnar epithelium that replaces the normal esophageal mucosa in patients with chronic gastroesophageal reflux. Because intestinal-type Barrett's metaplasia is the major risk factor for adenocarcinoma development, understanding the mechanisms that predispose the esophageal mucosa to malignant degeneration is clinically important. Glutathione s-transferase (GST)-pi belongs to a class of protective enzymes whose activity has been shown to be much lower in Barrett's metaplasia than in the normal esophagus, where this form of GST is predominant. In the studies described here, using immunocytochemical analysis, we observed higher levels of cytoplasmic GST-pi protein in normal esophageal mucosa than in Barrett's metaplasia. Using northern blot analysis, we also observed lower GST-pi mRNA levels in Barrett's metaplasia than in normal esophagus or adenocarcinomas from the same patients. Using as model systems three Barrett's adenocarcinoma cell lines and short-term organ culture of freshly resected normal esophagus and Barrett's metaplasia, dose-dependent induction of GST-pi mRNA was observed by using butylated hydroxyanisole and dexamethasone. GST-pi mRNA in Barrett's metaplasia was induced up to 2.5-fold with 60 microM butylated hydroxyanisole and nearly fivefold with 320 nM dexamethasone after 24 h. These studies demonstrate the ability to induce protective GST-pi in Barrett's metaplasia and may suggest a mechanism for future chemoprevention studies in patients with this type of epithelium, which is at high risk for malignant degeneration.

Needs and experiences of family caregivers during marrow transplantation.

PURPOSE/OBJECTIVES: To determine the information needs of family members of people undergoing marrow transplantation as well as their actions to meet those needs. DESIGN: Descriptive, cross-sectional, qualitative design. SETTING: Marrow transplant units in the Pacific Northwestern United States. SAMPLE: 19 adult family members of people who had undergone marrow transplants. METHODS: Researchers conducted four focus group interviews. Three served as data generation interviews, and the fourth served as the validation interview. A transcriptionist recorded the subjects' responses, which then were analyzed using constant comparative techniques. FINDINGS: Themes emerged from the data in five categories: (a) Preparing for Caregiving (seeking and acquiring health care, obtaining information and materials, and evaluating the validity of information), (b) Managing the Care (providing physical care, protecting, maintaining the patient's connection with life, and advocating), (c) Facing Challenges (personal and interpersonal stress, communication barriers with healthcare professionals, and healthcare system barriers), (d) Developing Supportive Strategies (community resources, personal and self-care resources, and healthcare system facilitators), and (e) Discovering Unanticipated Rewards and Benefits (personal growth and family cohesion). CONCLUSIONS: Family members, as well as patients undergoing marrow transplant, experience a unique set of information needs and demands as a result of this experience. However, these demands can be mitigated by actions that provide appropriate education strategies and foster a sense of caring and a nurturing way of interacting among the family, healthcare professionals, and the healthcare system. IMPLICATIONS FOR NURSING PRACTICE: Healthcare professionals need to acknowledge the caregiving role and actively involve and support the family caregiver throughout the transplant experience.

Understanding the caring relationship during marrow transplantation: family caregivers and healthcare professionals.

PURPOSE/OBJECTIVES: To understand the caring relationship that family caregivers of marrow transplant recipients wish to have with healthcare professionals within the context of Swanson's Caring Model. DATA SOURCES: Published articles, books, research results. DATA SYNTHESIS: This article describes how family caregivers of marrow transplant recipients experience caring relationships with healthcare providers within the context of the five caring processes identified by Swanson. Specific strategies for forming caring relationships with family caregivers are recommended. CONCLUSIONS: Healthcare professionals can enhance their caring relationship with family caregivers of marrow transplant recipients by embracing the five processes of Swanson's Caring Model: maintaining belief, knowing, being with, doing for, and enabling. IMPLICATIONS FOR NURSING PRACTICE: Strategies for forming caring relationships between healthcare professionals and family caregivers curing marrow transplantation include acknowledging family caregivers as individuals and providing clear expectations of the family caregiver role.

Educational interventions for family caregivers during marrow transplantation.

PURPOSE/OBJECTIVES: To make recommendations for the use of informational strategies and the development of resources for family caregivers of patients receiving marrow transplantation based on their identified informational needs. DATA SOURCES: Literature review and research that identifies informational needs of family caregivers of patients receiving marrow transplantation. DATA SYNTHESIS: Stetz, McDonald, and Compton described informational needs of family caregivers. The needs fell into five major categories: Preparing for Caregiving, Managing the Care, Facing Challenges, Developing Supportive Strategies, and Discovering Unanticipated Rewards and Benefits. CONCLUSIONS: Family members and close friends are assuming increasing responsibilities in providing care in the home for the patient during marrow transplantation. Much of this care occurs when the patient is being treated on an outpatient basis. The responsibilities are demanding and involve helping the patient access treatment, evaluate and deal with the stress of information, gain skills with complex procedures, manage symptoms at home, and communicate with healthcare providers. For this essential role, family caregivers need to be educated and supported by appropriate resources. IMPLICATIONS FOR NURSING PRACTICE: Family caregivers' need for information begins well before the patient's treatment is initiated at the selected center. Availability of accurate, specific written guidelines and personal contact is essential. Educational strategies that prepare family members before the caregiving skills are needed could include skill development, access to and use of resources, homecare management, decision making, and self-care.

Green fluorescent protein production in the cellular slime molds Polysphondylium pallidum and Dictyostelium discoideum.

The green fluorescent protein-encoding gene from Aequorea victoria has been cloned into several different transforming vectors and expressed in the cellular slime molds, Polysphondylium pallidum and Dictyostelium discoideum. We find that the protein is stable and non-toxic in both species, can be easily visualized in living and fixed specimens, and can be used to purify rare cells by fluorescence-activated cell sorting (FACS).

Development in one dimension: the rapid differentiation of Dictyostelium discoideum in glass capillaries.

When Dictyostelium discoideum cells are drawn into a fine glass capillary, they rapidly begin the first steps toward the formation of prestalk and prespore zones. Some of the events occur within a minute or two, whereas others follow later. The cells in the front segment are actively motile and those in the hind segment are passive. The volumes of the segments are proportional for different-sized cell masses, and those proportions are the same as those found in normal slugs. When the cells are stained with the vital dye neutral red, the anterior zone becomes darker simultaneously with the formation of the division line. Green fluorescent protein expressed from a stalk-specific promoter is synthesized mostly in the anterior end. Later, this capillary prestalk zone shows a sharp increase in alkaline phosphatase activity, which is known to be characteristic of prestalk cells.

Provisional practice: the nature of psychosocial bone marrow transplant nursing.

PURPOSE/OBJECTIVE: To develop an empirically based description of the key concepts guiding psychosocial nursing practice for the patient undergoing bone marrow transplantation (BMT). DESIGN: Retrospective, descriptive. SETTING: National Cancer Institute-designated comprehensive cancer center. SAMPLE: Medical records of 23 adults (11 males, 12 females; X age = 33 years) who have undergone BMT. METHODS: All psychosocial-related charting was extracted manually from the subjects' medical records and nursing care plans. Text was analyzed for content, coded, and sorted into 42 categories. The investigators wrote category definitions. MAIN RESEARCH VARIABLES: All present and past psychosocial problems that had been identified; psychosocial assessment; psychosocial-related admission and discharge teaching; and references to pain and sleep disturbances that included or implied psychosocial overlay. FINDINGS: Thematic categories were present in each of five core concepts: discovering the lived reality, managing the flow, emerging awareness, keeping watch, and behind closed doors. Synthesis of these concepts led to one encompassing concept--the provisional nature of psychosocial BMT nursing practice. CONCLUSIONS: The results provide empirical evidence that five key concepts exist within psychosocial BMT nursing and can be applied to a new paradigm of nursing care. IMPLICATIONS FOR NURSING PRACTICE: Provisional practice, as a new paradigm, requires further development. The five concepts derived from this study provide a useful framework for delineating significant psychosocial needs of patients and families and for designing tailored nursing therapeutics. The study suggests that preparation for practicing BMT nursing should include education in specific psychosocial strategies, documentation, and coping with the personal implications of practice in this setting. The study raises important issues in psychosocial care requiring further clarification and elucidation. These center around the nature of uncertainty, caring, and social support in BMT.

Long-term treatment of alcoholic liver disease with propylthiouracil. Part 2: Influence of drop-out rates and of continued alcohol consumption in a clinical trial.

Although propylthiouracil has previously been shown to reduce the risk of mortality in alcoholic liver disease by 60%, generalized use of propylthiouracil for this condition has not occurred. Additional data are therefore presented on four aspects to provide a better assessment of its therapeutic effectiveness. First, the characteristics and the prognosis of dropouts were virtually identical in both the drug and placebo groups. Also the methodology and analysis employed, were designed to control for dropouts, thus providing an accurate interpretation of the outcome. Secondly, since 97% of the patients continued to drink, abstinence was not a precondition for the beneficial effect of propylthiouracil. However, the beneficial effect was observed most clearly in those patients who continued to drink at lower levels, whereas lower level drinking per se did not afford protection in placebo patients. Thirdly, serious side effects or clinical hypothyroidism occurred extremely rarely in these patients, many of whom have now received propylthiouracil for over 4 years. Fourthly, we discuss why the outcome in long-term clinical trials in alcoholic liver disease cannot be compared with effects observed in clinical trials lasting only a few weeks. Journal of Hepatology.