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Since December 2019, the global pandemic caused by the highly infectious novel coronavirus 2019-nCoV (COVID-19) has been rapidly spreading. As of April 2020, the outbreak has spread to over 210 countries, with over 2,400,000 confirmed cases and over 170,000 deaths. 1 COVID-19 causes a severe pneumonia characterized by fever, cough and shortness of breath. Similar coronavirus outbreaks have occurred in the past causing severe pneumonia like COVID-19, most recently, severe acute respiratory syndrome coronavirus (SARS-CoV) and middle east respiratory syndrome coronavirus (MERS-CoV). However, over time, SARS-CoV and MERS-CoV were shown to cause extrapulmonary signs and symptoms including hepatitis, acute renal failure, encephalitis, myositis and gastroenteritis. Similarly, sporadic reports of COVID-19 related extrapulmonary manifestations emerge. Unfortunately, there is no comprehensive summary of the multiorgan manifestations of COVID-19, making it difficult for clinicians to quickly educate themselves about this highly contagious and deadly pathogen. What is more, is that SARS-CoV and MERS-CoV are the closest humanity has come to combating something similar to COVID-19, however, there exists no comparison between the manifestations of any of these novel coronaviruses. In this review, we summarize the current knowledge of the manifestations of the novel coronaviruses SARS-CoV, MERS-CoV and COVID-19, with a particular focus on the latter, and highlight their differences and similarities., (Copyright © 2020 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.)

Aim: The burden on nursing staff in Germany is continuously increasing. Specific health promotion programs can help to reduce this burden. To develop such programs, the concrete needs of nurses for instructions and training have to be determined. The aim of the project "PFLEGEprevent" (funded by the Bayerisches Staatsministerium für Gesundheit und Pflege) is to develop a prevention program that focuses on the special needs of the nursing staff. The project is structured in four subprojects: (1) identification of needs, workloads and occupational challenges of caregivers and the scientific evidence (national expert survey and systematic literature review), (2) development of a specific prevention program, (3) implementation and evaluation of the prevention program (Intervention study; DRKS00012870), (4) development of a manual to instruct the implementation of the prevention program (transferability)., Method: In subproject (1) the current subjective work ability of the nursing staff in Germany and their need for preventive offers were revealed by a national expert survey. The survey was directed to nurses in outpatient, semi-inpatient and inpatient care facilities and clinics. In addition to personal information and questions on current work and organization, the subjective work ability of the nurses (based on the Work Ability Index, WAI) and the need for preventive offers were queried., Results: A total of 1381 questionnaires were analysed. The majority of surveyed healthcare staff was female (81%). On average they were 40.1 years old (SD=12.0) and had worked in the care sector for 16.3 years (SD=10.9). The mean of the WAI score was 37.2 (SD=7.1) points, which indicates a good work ability. However, there were differences in work ability between types of institution and number of years in job. The care staff needed instructions mainly on the topics of stress, communication and teamwork. Active training was required for relaxation, back muscle training and strengthening. 75% of respondents said they would participate in a 5-day prevention program., Competing Interests: Die Autoren geben an, dass kein Interessenkonflikt besteht., (© Georg Thieme Verlag KG Stuttgart · New York.)

DNA can be preserved in marine and freshwater sediments both in bulk sediment and in intact, viable resting stages. Here, we assess the potential for combined use of ancient, environmental, DNA and timeseries of resurrected long-term dormant organisms, to reconstruct trophic interactions and evolutionary adaptation to changing environments. These new methods, coupled with independent evidence of biotic and abiotic forcing factors, can provide a holistic view of past ecosystems beyond that offered by standard palaeoecology, help us assess implications of ecological and molecular change for contemporary ecosystem functioning and services, and improve our ability to predict adaptation to environmental stress.

Knowledge of the molecular basis of phenotypic responses to environmental cues is key to understanding the process of adaptation. Insights to adaptation at an evolutionary time scale can be gained by observing organismal responses before and after a shift in environmental conditions, but such observations can rarely be made. Using the ecological and genomic model Daphnia, we linked transcriptomic responses and phosphorus (P)-related phenotypic traits under high and low P availability. We mapped weighted gene coexpression networks to traits previously assessed in resurrected ancient (600 years old) and modern Daphnia pulicaria from a lake with a historic shift in P-enrichment. Subsequently, we assessed evolutionary conservation or divergence in transcriptional networks of the same isolates. We discovered highly preserved gene networks shared between ancient genotypes and their modern descendants, but also detected clear evidence of transcriptional divergence between these evolutionarily separated genotypes. Our study highlights that phenotypic evolution is a result of molecular fine-tuning on different layers ranging from basic cellular responses to higher order phenotypes. In a broader context, these findings advance our understanding how populations are able to persist throughout major environmental shifts., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Atrioventricular (AV) block is usually due to infranodal disease and associated with a wide QRS complex; such patients often progress to complete AV block and pacemaker dependency. Uncommonly, infranodal AV block can occur within the His bundle with a narrow QRS complex., Objectives: The aims of this study were to define clinical/echocardiographic characteristics of patients with AV block within the His bundle and report progression to pacemaker dependency., Methods: We retrospectively identified patients with narrow QRS complexes and documented intra-His delay or block at electrophysiology study (group A) or with electrocardiogram-documented Mobitz II AV block/paroxysmal AV block (group B). Clinical, electrophysiological, and echocardiographic variables at presentation and pacemaker parameters at the last follow-up visit were evaluated., Results: Twenty-seven patients (19 women) were identified (mean age 64 ± 13 years; range, 38-85 years). Four patients who had <1 month of follow-up were excluded. There were 12 patients in group A and 11 in group B; 21 of 23 presented with syncope/presyncope. All patients received pacemakers: 8 single chamber and 15 dual chamber. After a median follow-up of 6.4 years, the median percentage of ventricular pacing was 1% (interquartile range 0%-4.66%). One patient developed true pacemaker dependency. Aortic and/or mitral annular calcification was present in 13 of 22 patients with available echocardiograms., Conclusion: Patients who present with syncope and narrow QRS complexes with intra-His delay or Mobitz II paroxysmal AV block with narrow QRS complexes rarely progress to pacemaker dependency and require infrequent pacing. This entity is more common in women, with a higher prevalence of aortic and/or mitral annular calcification. If confirmed by additional studies, single-chamber pacemaker may be sufficient., (Copyright © 2018 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.)

Paleolimnologists have utilized lake sediment records to understand historical lake and landscape development, timing and magnitude of environmental change at lake, watershed, regional and global scales, and as historical datasets to target watershed and lake management. Resurrection ecologists have long recognized lake sediments as sources of viable propagules ("seed or egg banks") with which to explore questions of community ecology, ecological response, and evolutionary ecology. Most researchers consider Daphnia as the primary model organism in these efforts, but many other aquatic biota, from viruses to macrophytes, similarly produce viable propagules that are incorporated in the sediment record but have been underutilized in resurrection ecology. The common goals shared by these two disciplines have led to mutualistic and synergistic collaborations-a development that must be encouraged to expand. We give an overview of the achievements of paleolimnology and the reconstruction of environmental history of lakes, review the untapped diversity of aquatic organisms that produce dormant propagules, compare Daphnia as a model of resurrection ecology with other organisms amenable to resurrection studies, especially diatoms, and consider new research directions that represent the nexus of these two fields.

This perspective provides an overview to the Special Issue on Resurrection Ecology (RE). It summarizes the contributions to this Special Issue, and provides background information and future prospects for the use of RE in both basic and applied evolutionary studies.

Understanding the colonisation process in zooplankton is crucial for successful restoration of aquatic ecosystems. Here, we analyzed the clonal and genetic structure of the cyclical parthenogenetic rotifer Brachionus plicatilis by following populations established in new temporary ponds during the first three hydroperiods. Rotifer populations established rapidly after first flooding, although colonisation was ongoing throughout the study. Multilocus genotypes from 7 microsatellite loci suggested that most populations (10 of 14) were founded by few clones. The exception was one of the four populations that persisted throughout the studied hydroperiods, where high genetic diversity in the first hydroperiod suggested colonisation from a historical egg bank, and no increase in allelic diversity was detected with time. In contrast, in another of these four populations, we observed a progressive increase of allelic diversity. This population became less differentiated from the other populations suggesting effective gene flow soon after its foundation. Allelic diversity and richness remained low in the remaining two, more isolated, populations, suggesting little gene flow. Our results highlight the complexity of colonisation dynamics, with evidence for persistent founder effects in some ponds, but not in others, and with early immigration both from external source populations, and from residual, historical diapausing egg banks.

Understanding the evolutionary consequences of the green revolution, particularly in wild populations, is an important frontier in contemporary biology. Because human impacts have occurred at varying magnitudes or time periods depending on the study ecosystem, evolutionary histories may vary considerably among populations. Paleogenetics in conjunction with paleolimnology enable us to associate microevolutionary dynamics with detailed information on environmental change. We used this approach to reconstruct changes in the temporal population genetic structure of the keystone zooplankton grazer, Daphnia pulicaria, using dormant eggs extracted from sediments in two Minnesota lakes (South Center, Hill). The extent of agriculture and human population density in the catchment of these lakes has differed markedly since European settlement in the late 19th century and is reflected in their environmental histories reconstructed here. The reconstructed environments of these two lakes differed strongly in terms of environmental stability and their associated patterns of Daphnia population structure. We detected long periods of stability in population structure and environmental conditions in South Center Lake that were followed by a dramatic temporal shift in population genetic structure after the onset of European settlement and industrialized agriculture in its watershed. In particular, we noted a 24.3-fold increase in phosphorus (P) flux between pre-European and modern sediment P accumulation rates (AR) in this lake. In contrast, no such shifts were detected in Hill Lake, where the watershed was not as impacted by European settlement and rates of change were less directional with a much smaller increase in sediment P AR (2.3-fold). We identify direct and indirect effects of eutrophication proxies on genetic structure in these lake populations and demonstrate the power of using this approach in understanding the consequences of anthropogenic environmental change on natural populations throughout historic time periods., (© 2016 John Wiley & Sons Ltd.)

Background: Chronic psychological distress appears to have increased in recent years, mainly among the working population. The data available indicate that mental and behavioral disorders, including burnout syndrome, represent not only a personal problem for those afflicted, but also a serious public health issue. This study aimed at evaluating the effects of an outpatient burnout prevention program in a mono-center health resort setting., Methods: Adults experiencing an above-average level of stress and thus being at an increased risk of burnout were randomized either to the intervention group (IG) or the waiting control group (WG). The 3-week program included stress management intervention, relaxation, physical exercise and moor applications. The primary outcome was change in perceived stress (PSQ) at 6 months post-intervention. Secondary outcomes included burnout symptoms, well-being, health status, psychological symptoms, back pain, and number of sick days. Participants were examined at baseline, post-intervention (3 weeks) and after 1, 3 and 6 months., Results: Data from 88 adults (IG=43; WG=45) were available for (per protocol) analysis (mean age: 50.85; 76.1% female). Participants in the IG experienced significant immediate improvement in all outcome measures, which declined somewhat during the first three months post-intervention and then remained stable for at least another three months. Those in the WG did not experience substantial change across time. For the 109 randomized persons, results for PSQ were confirmed in an intention-to-treat analysis with missing values replaced by last observation carried forward (between-group ANCOVA for PSQScore at 6 months, parameter estimator for the group: -20.57; 95% CI: [-26.09; -15.04]). Large effect sizes (Cohen's d for PSQ: 1.09-1.72) indicate the superiority of the intervention., Conclusion: The program proved to be effective in reducing perceived stress, emotional exhaustion and other targets. Future research should examine the long-term impact of the program and the effect of occasional refresher training.

Objective: The aim of this study was to investigate the reasons of the insured in Germany for taking cure regimes in foreign European countries., Methods: Health insurances conducted a survey including insurance holders in Germany who had taken a cure regime in a foreign European country in 2012 or 2011. In a paper-pencil interview, the sample was asked why they performed the cure regime abroad, how satisfied they were regarding the quality of the health professionals and the treatment, respectively, how long a positive effect lasted, where they spent their cure regime and about their plans regarding a future treatment in a health resort., Results: In total, 443 insurance holders (60.7% female, mean age 68 years) were included. Price-performance ratio of board, lodging and treatment (75.6%), the health resorts' reputation (64.5%) and recommendations of the personal environment (56.1%) were the main reasons for taking a cure regime abroad. Most of the participants were satisfied with the quality of the health professionals and the health resort; two thirds rated the effect as lasting longer than 3 months. The study population predominantly took services of health resorts located in the Czech Republic, in Hungary and in Poland. For taking a cure in the future, most of the study participants (77%) preferred health resorts abroad, however, a gender-specific analysis revealed this trend to be more pronounced in the male study population., Conclusion: Based on the results of the survey one might suggest that cost factors along with high level of satisfaction regarding the quality of treatments provided at the health resorts abroad, positively influence the decision for taking a cure regime in a foreign country. In the sample of this study, the decision is rarely based on the health insurances' or the health resorts' promotion activity, respectively. However, to conclude on the total population, requires data of a representative sample., (© Georg Thieme Verlag KG Stuttgart · New York.)

Whole-chamber mapping using a 64-pole basket catheter (BC) has become a featured approach for the analysis of excitation patterns during atrial fibrillation. A flexible catheter design avoids perforation but may lead to spline bunching and influence coverage. We aim to quantify the catheter deformation and endocardial coverage in clinical situations and study the effect of catheter size and electrode arrangement using an in silico basket model. Atrial coverage and spline separation were evaluated quantitatively in an ensemble of clinical measurements. A computational model of the BC was implemented including an algorithm to adapt its shape to the atrial anatomy. Two clinically relevant mapping positions in each atrium were assessed in both clinical and simulated data. The simulation environment allowed varying both BC size and electrode arrangement. Results showed that interspline distances of more than 20 mm are common, leading to a coverage of less than 50% of the left atrial (LA) surface. In an ideal in silico scenario with variable catheter designs, a maximum coverage of 65% could be reached. As spline bunching and insufficient coverage can hardly be avoided, this has to be taken into account for interpretation of excitation patterns and development of new panoramic mapping techniques., Competing Interests: The authors declare that there is no conflict of interests regarding the publication of this paper.

Health resort medicine (HRM; in German: Kurortmedizin) is a field of medicine with long-lasting tradition in several European countries. A number of systematic reviews have shown the effectiveness of HRM in musculoskeletal conditions. Reviews focusing on the effectiveness of HRM in non-musculoskeletal disorders are rare. This systematic review aims to provide an overview about all types of health resort treatments applied in non-musculoskeletal conditions, to summarize evidence for its effectiveness and to assess the quality of published studies. MEDLINE, Web of Knowledge and Embase were searched for articles published between January 2002 and December 2013. We used a broad search strategy in order to find studies investigating the effects of HRM in non-musculoskeletal disorders. Two authors independently extracted data and assessed quality using the Effective Public Health Practice Project Quality Assessment Tool for Quantitative Studies (EPHPP-QAT). Forty-one studies (19 of them with control group) from eight countries examining the efficacy of various forms of spa treatment for 12 disease groups were included. The studies are markedly heterogeneous regarding study design, population and treatment. HRM treatment is associated with clinical improvement in diseases of the skin, respiratory, circulatory, digestive and nervous system among others. However, small samples, the lack of control groups and an insufficient follow-up often limit the generated evidence. The scientific literature of the last decade has shown that a number of non-musculoskeletal disorders are treated with different kinds of HRM. The challenge for the future will be to carry out thoroughly designed studies in larger patient populations to corroborate the impact of HRM treatment on non-musculoskeletal disorders.

Little is known about the role of transcriptomic changes in driving phenotypic evolution in natural populations, particularly in response to anthropogenic environmental change. Previous analyses of Daphnia genotypes separated by centuries of evolution in a lake using methods in resurrection ecology revealed striking genetic and phenotypic shifts that were highly correlated with anthropogenic environmental change, specifically phosphorus (P)-driven nutrient enrichment (i.e. eutrophication). Here, we compared the transcriptomes of two ancient (~700-year-old) and two modern (~10-year-old) genotypes in historic (low P) and contemporary (high P) environmental conditions using microarrays. We found considerable transcriptomic variation between 'ancient' and 'modern' genotypes in both treatments, with stressful (low P) conditions eliciting differential expression (DE) of a larger number of genes. Further, more genes were DE between 'ancient' and 'modern' genotypes than within these groups. Expression patterns of individual genes differed greatly among genotypes, suggesting that different transcriptomic responses can result in similar phenotypes. While this confounded patterns between 'ancient' and 'modern' genotypes at the gene level, patterns were discernible at the functional level: annotation of DE genes revealed particular enrichment of genes involved in metabolic pathways in response to P-treatments. Analyses of gene families suggested significant DE in pathways already known to be important in dealing with P-limitation in Daphnia as well as in other organisms. Such observations on genotypes of a single natural population, separated by hundreds of years of evolution in contrasting environmental conditions before and during anthropogenic environmental changes, highlight the important role of transcriptional mechanisms in the evolutionary responses of populations., (© 2014 John Wiley & Sons Ltd.)

With medical advances, more individuals with cerebral palsy (CP) syndromes who reside in developed countries are surviving to adolescence and adulthood. However, there continues to be a paucity of research examining long-term health, functional activities, and participatory outcomes over their life-course. This article reviews the current literature assessing adult outcomes for individuals with CP within the framework of the International Classification of Functioning (ICF), Disability, and Health model of enablement. Preliminary data over the last decade indicate that among adults with cerebral palsy without intellectual disability, 60-80% completed high school, 14-25% completed college, up to 61% were living independently in the community, 25-55% were competitively employed, and 14-28% were involved in long term relationships with partners or had established families. These outcomes occurred with biomedical advances in the management of spasticity, deformity, and medical co-morbidities, as well as with concurrent policy initiatives to increase access to a continuum of habilitative and education services. Although we have incomplete population data to inform comprehensive life-course planning, there are opportunities to create clinical and translational community networks with improved measures of functioning and participation that can better inform us about the factors influencing lifespan development of people with CP., (Copyright © 2013 Wiley Periodicals, Inc., a Wiley company.)

The magnitude of community-wide dispersal is central to metacommunity models, yet dispersal is notoriously difficult to quantify in passive and cryptic dispersers such as many freshwater invertebrates. By overcoming the problem of quantifying dispersal rates, colonization rates into new habitats can provide a useful estimate of the magnitude of effective dispersal. Here we study the influence of spatial and local processes on colonization rates into new ponds that indicate differential dispersal limitation of major zooplankton taxa, with important implications for metacommunity dynamics. We identify regional and local factors that affect zooplankton colonization rates and spatial patterns in a large-scale experimental system. Our study differs from others in the unique setup of the experimental pond area by which we were able to test spatial and environmental variables at a large spatial scale. We quantified colonization rates separately for the Copepoda, Cladocera and Rotifera from samples collected over a period of 21 months in 48 newly constructed temporary ponds of 0.18-2.95 ha distributed in a restored wetland area of 2,700 ha in Doñana National Park, Southern Spain. Species richness upon initial sampling of new ponds was about one third of that in reference ponds, although the rate of detection of new species from thereon were not significantly different, probably owing to high turnover in the dynamic, temporary reference ponds. Environmental heterogeneity had no detectable effect on colonization rates in new ponds. In contrast, connectivity, space (based on latitude and longitude) and surface area were key determinants of colonization rates for copepods and cladocerans. This suggests dispersal limitation in cladocerans and copepods, but not in rotifers, possibly due to differences in propagule size and abundance.

Interleukin-10 (IL-10) is a potent anti-inflammatory cytokine, with therapeutic applications in inflammatory bowel disease. For the in situ delivery of IL-10 by Escherichia coli as carrier chassis, a modified transporter was designed with the ability to secrete biologically active IL-10. De novo DNA synthesis comprised a 561-bp fragment encoding the signal sequence of the E. coli outer membrane protein F fused in frame to an E. coli codon-optimized mature human IL-10 gene under control of a T7 promoter. The construct was overexpressed in E. coli laboratory strains, E. coli BL21 (DE3) and E. coli MDS42:T7. The mean concentrations of human IL-10 in the periplasm and culture supernatant of E. coli BL21 (DE3) were 355.8 ± 86.3 and 5.7 ± 1.7 ng/ml, respectively. The molecular mass of the recombinant E. coli-derived human IL-10 was 19 kDa, while under non-reducing conditions the native IL-10 dimer could be demonstrated. Reduction of tumor necrosis factor-α secretion in lipopolysaccharide-stimulated mouse macrophages and detection of the activated form of the transcription factor signal transducer and activator of transcription protein 3 proved the biological activity of the bacteria-produced human IL-10., (Copyright © 2012 S. Karger AG, Basel.)

Background: Copepods outnumber every other multicellular animal group. They are critical components of the world's freshwater and marine ecosystems, sensitive indicators of local and global climate change, key ecosystem service providers, parasites and predators of economically important aquatic animals and potential vectors of waterborne disease. Copepods sustain the world fisheries that nourish and support human populations. Although genomic tools have transformed many areas of biological and biomedical research, their power to elucidate aspects of the biology, behavior and ecology of copepods has only recently begun to be exploited., Discussion: The extraordinary biological and ecological diversity of the subclass Copepoda provides both unique advantages for addressing key problems in aquatic systems and formidable challenges for developing a focused genomics strategy. This article provides an overview of genomic studies of copepods and discusses strategies for using genomics tools to address key questions at levels extending from individuals to ecosystems. Genomics can, for instance, help to decipher patterns of genome evolution such as those that occur during transitions from free living to symbiotic and parasitic lifestyles and can assist in the identification of genetic mechanisms and accompanying physiological changes associated with adaptation to new or physiologically challenging environments. The adaptive significance of the diversity in genome size and unique mechanisms of genome reorganization during development could similarly be explored. Genome-wide and EST studies of parasitic copepods of salmon and large EST studies of selected free-living copepods have demonstrated the potential utility of modern genomics approaches for the study of copepods and have generated resources such as EST libraries, shotgun genome sequences, BAC libraries, genome maps and inbred lines that will be invaluable in assisting further efforts to provide genomics tools for copepods., Summary: Genomics research on copepods is needed to extend our exploration and characterization of their fundamental biological traits, so that we can better understand how copepods function and interact in diverse environments. Availability of large scale genomics resources will also open doors to a wide range of systems biology type studies that view the organism as the fundamental system in which to address key questions in ecology and evolution.

Objective: The aim of this trial was to investigate the effectiveness of a worksite intervention using kettlebell training to improve musculoskeletal and cardiovascular health., Methods: This single-blind randomized controlled trial involved 40 adults from occupations with a high prevalence of reported musculoskeletal pain symptoms (mean age 44 years, body mass index 23 kg/m², 85% women, with pain intensity of the neck/shoulders 3.5 and of the low back 2.8 on a scale of 0-10). A blinded assessor took measures at baseline and follow-up. Participants were randomly assigned to training--consisting of ballistic full-body kettlebell exercise 3 times per week for 8 weeks--or a control group. The main outcome measures were pain intensity of the neck/shoulders and low back, isometric muscle strength, and aerobic fitness., Results: Compared with the control group, pain intensity of the neck/shoulders decreased 2.1 points [95% confidence interval (95% CI) -3.7- -0.4] and pain intensity of the low back decreased 1.4 points (95% CI -2.7- -0.02) in the training group. Compared with the control group, the training group increased muscle strength of the trunk extensors (P<0.001), but not of the trunk flexors and shoulders. Aerobic fitness remained unchanged., Conclusions: Worksite intervention using kettlebell training reduces pain in the neck/shoulders and low back and improves muscle strength of the low back among adults from occupations with a high prevalence of reported musculoskeletal pain symptoms. This type of training does not appear to improve aerobic fitness.

Population genetic surveys approximately 25 years apart examined the distribution and abundance of asexual clones of the freshwater zooplankter Daphnia pulex complex in rock pools near Churchill, Manitoba, Canada. In 1984-1985, melanic members of this species complex were present in 131 rock pools at this site, but were only detected in 90 of these pools in 2007-2008. Allozymic surveys conducted during these two time periods revealed that 59 per cent of these populations showed unchanged clonal composition. Total clonal replacement occurred in 8 per cent of the populations, while the others (33%) included a mixture of 'resident' clones and new 'colonists'. We discuss these changes in light of shifts in biotic and abiotic factors. We also discuss the use of rock pool habitats as 'sentinel' systems for examining long-term environmental changes in the ecological genetics of aquatic organisms in the Arctic.

Highly reduced E. coli strains, MDS40, MDS41, and MDS42, lacking approximately 15% of the genome, were grown to high cell densities to test their ability to produce a recombinant protein with high yields. These strains lack all transposons and insertion sequences, cryptic prophage and many genes of unknown function. In addition to improving genetic stability, these deletions may reduce the biosynthetic requirements of the cell potentially allowing more efficient production of recombinant protein. Basic growth parameters and the ability of the strains to produce chloramphenicol acetyltransferase (CAT) under high cell density, batch cultivation were assessed. Although growth rate and recombinant protein production of the reduced genome strains are comparable to the parental MG1655 strain, the reduced genome strains were found to accumulate significant amounts of acetate in the medium at the expense of additional biomass. A number of hypotheses were examined to explain the accumulation of acetate, including oxygen limitation, carbon flux imbalance, and metabolic activity of the recombinant protein. Use of a non-catalytic CAT variant identified the recombinant protein activity as the source of this phenomenon; implications for the metabolic efficiency of the reduced genome strains are discussed.

With the use of synthetic biology, we reduced the Escherichia coli K-12 genome by making planned, precise deletions. The multiple-deletion series (MDS) strains, with genome reductions up to 15%, were designed by identifying nonessential genes and sequences for elimination, including recombinogenic or mobile DNA and cryptic virulence genes, while preserving good growth profiles and protein production. Genome reduction also led to unanticipated beneficial properties: high electroporation efficiency and accurate propagation of recombinant genes and plasmids that were unstable in other strains. Eradication of stress-induced transposition evidently stabilized the MDS genomes and provided some of the new properties.

A high-throughput method has been developed for the systematic mutagenesis of the Escherichia coli genome. The system is based on in vitro transposition of a modified Tn5 element, the Sce-poson, into linear fragments of each open reading frame. The transposon introduces both positive (kanamycin resistance) and negative (I-SceI recognition site) selectable markers for isolation of mutants and subsequent allele replacement, respectively. Reaction products are then introduced into the genome by homologous recombination via the lambdaRed proteins. The method has yielded insertion alleles for 1976 genes during a first pass through the genome including, unexpectedly, a number of known and putative essential genes. Sce-poson insertions can be easily replaced by markerless mutations by using the I-SceI homing endonuclease to select against retention of the transposon as demonstrated by the substitution of amber and/or in-frame deletions in six different genes. This allows a Sce-poson-containing gene to be specifically targeted for either designed or random modifications, as well as permitting the stepwise engineering of strains with multiple mutations. The promiscuous nature of Tn5 transposition also enables a targeted gene to be dissected by using randomly inserted Sce-posons as shown by a lacZ allelic series. Finally, assessment of the insertion sites by an iterative weighted matrix algorithm reveals that these hyperactive Tn5 complexes generally recognize a highly degenerate asymmetric motif on one end of the target site helping to explain the randomness of Tn5 transposition.

Cotton fibers are single-celled seed trichomes of major economic importance. Factors that regulate the rate and duration of cell expansion control fiber morphology and important agronomic traits. For genetic characterization of rapid cell elongation in cotton fibers, approximately 14,000 unique genes were assembled from 46,603 expressed sequence tags (ESTs) from developmentally staged fiber cDNAs of a cultivated diploid species ( Gossypium arboreum L.). Conservatively, the fiber transcriptome represents 35-40% of the genes in the cotton genome. In silico expression analysis revealed that rapidly elongating fiber cells exhibit significant metabolic activity, with the bulk of gene transcripts, represented by three major functional groups - cell wall structure and biogenesis, the cytoskeleton and energy/carbohydrate metabolism. Oligonucleotide microarrays revealed dynamic changes in gene expression between primary and secondary cell wall biogenesis showing that fiber genes in the dbEST are highly stage-specific for cell expansion - a conclusion supported by the absence of known secondary cell wall-specific genes from our fiber dbEST. During the developmental switch from primary to secondary cell wall syntheses, 2553 "expansion-associated" fiber genes are significantly down regulated. Genes (81) significantly up-regulated during secondary cell wall synthesis are involved in cell wall biogenesis and energy/carbohydrate metabolism, which is consistent with the stage of cellulose synthesis during secondary cell wall modification in developing fibers. This work provides the first in-depth view of the genetic complexity of the transcriptome of an expanding cell, and lays the groundwork for studying fundamental biological processes in plant biology with applications in agricultural biotechnology.

Although the time for contrast material to fill the epicardial artery in the setting of acute coronary syndromes has been studied extensively, the time for contrast material to fill the myocardium has not been evaluated. We compared differences in myocardial contrast material transit among patients with unstable angina pectoris/non-ST-elevation acute myocardial infarction (UAP/NSTEAMI) with patients with ST-elevation acute myocardial infarction (STEAMI). The time it took for contrast material to first appear and to arrive at peak intensity in the myocardium was compared in 224 patients with STEAMI enrolled in the LIMIT-AMI study versus 430 patients with UAP/NSTEAMI enrolled in the TACTICS-TIMI 18 trial. In patients with STEAMI, there was a delay in both the time for contrast material to first enter the myocardium (5,619 +/- 1,789 vs 4,663 +/- 1,626 ms, p <0.0001) and the time from entrance to peak blush intensity (2,387 +/- 1,359 vs 1,959 +/- 1,244 ms, p = 0.003) compared with patients with UAP/NSTEAMI. STEAMI remained significantly associated with impaired entrance of contrast material into the myocardium (p <0.0001) in a multivariate model controlling for known correlates of impaired epicardial flow (presence of thrombus, percent diameter stenosis, left anterior descending artery location, and contrast material inflow in the epicardial artery [corrected TIMI frame count]). The time for contrast material to enter the myocardium is impaired to a greater degree in STEAMI compared with UAP/NSTEAMI, even after adjusting for other variables known to delay flow in the epicardial artery. These data provide insight into potential mechanistic differences between these 2 clinical syndromes.

We have constructed a bacterial artificial chromosome (BAC) library for a European honey bee strain using the cloning enzyme HindIII in order to develop resources for structural genomics research. The library contains 36,864 clones (ninety-six 384-well plates). A random sampling of 247 clones indicated an average insert size of 113 kb (range = 27 to 213 kb) and 2% empty vectors. Based on an estimated genome size of 270 Mb, this library provides approximately 15 haploid genome equivalents, allowing >99% probability of recovering any specific sequence of interest. High-density colony filters were gridded robotically using a Genetix Q-BOT in a 4 x 4 double-spotted array on 22.5-cm2 filters. Screening of the library with four mapped honey bee genomic clones and two bee cDNA probes identified an average of 21 positive signals per probe, with a range of 7-38 positive signals per probe. An additional screening was performed with nine aphid gene fragments and one Drosophila gene fragment resulting in seven of the nine aphid probes and the Drosophila probe producing positive signals with a range of 1 to 122 positive signals per probe (average of 45). To evaluate the utility of the library for sequence tagged connector analysis, 1152 BAC clones were end sequenced in both forward and reverse directions, giving a total of 2061 successful reads of high quality. End sequences were queried against SWISS-PROT, insect genomic sequence GSS, insect EST, and insect transposable element databases. Results in spreadsheet format from these searches are publicly available at the Clemson University Genomics Institute (CUGI) website in a searchable format (http://www.genome.clemson.edu/projects/stc/bee/AM&#95;&#95;Ba/).

Background: Stenting has been shown to improve lumen diameters and thereby improve epicardial blood flow, but the impact of stent placement on tissue level perfusion has not been well characterized., Methods: Data were drawn from the LIMIT trial of rhuMAb CD18 (anti WBC antibody) in acute myocardial infarction (AMI). Adjunctive/rescue stenting was performed at the discretion of the investigator. The TIMI Myocardial Perfusion Grade (TMPG) was assessed and digital subtraction angiography (DSA) was used to quantify brightness of the myocardial blush., Results: TIMI 3 flow was 54.2% (64/118) before stent placement, and improved to 87.2% (102/117, p < 0.001) following stent placement. Likewise, Corrected TIMI Frame Counts (CTFCs) improved from medians of 37.6 to 21 (p < 0.001). By DSA, the rate of growth in brightness also tended to be greater after stenting (2.3 +/- 0.4 Gray/sec, n = 54 vs 3.1 +/- 0.3, n = 54, p = 0.07). The incidence of TMPG 0 decreased following stent placement (25.2% (29/118) vs 14.3% (16/118), p = 0.03) and the incidence of a stain in the myocardium (TMPG 1) increased (13.5% (16/118) vs 28.6% (34/118), p = 0.004)., Conclusion: Adjunctive stenting following thrombolytic administration in AMI improves epicardial TIMI 3 flow and TIMI frame counts as well as dye inflow into the myocardium: TMPG 0 is reduced and myocardial blush measured quantitatively by DSA tends to be brighter. However, more TMPG 1 or dye staining was present on next injection, suggesting dye outflow may be impaired.

Rice was chosen as a model organism for genome sequencing because of its economic importance, small genome size, and syntenic relationship with other cereal species. We have constructed a bacterial artificial chromosome fingerprint-based physical map of the rice genome to facilitate the whole-genome sequencing of rice. Most of the rice genome ( approximately 90.6%) was anchored genetically by overgo hybridization, DNA gel blot hybridization, and in silico anchoring. Genome sequencing data also were integrated into the rice physical map. Comparison of the genetic and physical maps reveals that recombination is suppressed severely in centromeric regions as well as on the short arms of chromosomes 4 and 10. This integrated high-resolution physical map of the rice genome will greatly facilitate whole-genome sequencing by helping to identify a minimum tiling path of clones to sequence. Furthermore, the physical map will aid map-based cloning of agronomically important genes and will provide an important tool for the comparative analysis of grass genomes.

Utilizing improved methods, two bacterial artificial chromosome (BAC) libraries were constructed for the multidisease-resistant line of melon MR-1. The HindIII library consists of 177 microtiter plates in a 384-well format, while the EcoRI library consists of 222 microtiter plates. Approximately 95.6% of the HindIII library clones contain nuclear DNA inserts with an average size of 118 kb, providing a coverage of 15.4 genome equivalents. Similarly, 96% of the EcoRI library clones contain nuclear DNA inserts with an average size of 114 kb, providing a coverage of 18.7 genome equivalents. Both libraries were evaluated for contamination with high-copy vector, empty pIndigoBac536 vector, and organellar DNA sequences. High-density filters were screened with two genetic markers FM and AM that co-segregate with Fom-2, a gene conferring resistance to races 0 and 1 of Fusarium wilt. Fourteen and 18 candidate BAC clones were identified for the FM and AM probes, respectively, from the HindIII library, while 34 were identified for the AM probe from filters A, B, and C of the EcoRI library.

Rice is an important food crop and a model plant for other cereal genomes. The Clemson University Genomics Institute framework project, begun two years ago in anticipation of the now ongoing international effort to sequence the rice genome, is nearing completion. Two bacterial artificial chromosome (BAC) libraries have been constructed from the Oryza sativa cultivar Nipponbare. Over 100,000 BAC end sequences have been generated from these libraries and, at a current total of 28 Mbp, represent 6.5% of the total rice genome sequence. This sequence information has allowed us to draw first conclusions about unique and redundant rice genomic sequences. In addition, more than 60,000 clones (19 genome equivalents) have been successfully fingerprinted and assembled into contigs using FPC software. Many of these contigs have been anchored to the rice chromosomes using a variety of techniques. Hybridization experiments have shown these contigs to be very robust. Contig assembly and hybridization experiments have revealed some surprising insights into the organization of the rice genome, which will have significant repercussions for the sequencing effort. Integration of BAC end sequence data with anchored contig information has provided unexpected revelations on sequence organization at the chromosomal level.

Objective: To identify health perceptions and practices and non-adherence to therapy among tuberculosis patients., Material and Methods: Qualitative research work consisting of 11 group interviews with 62 tuberculosis patients during 1997-1998 in the Central, Highlands, and Border Regions of Chiapas, Mexico., Results: Perceived causes of tuberculosis included contagion via food utensils, excess work, malnutrition, and cold, as well as other causes unrelated to person-to-person contagion. The resulting incapability to work resulted in an economic crisis for both the patients and their family members. As a result of the social stigma imposed by the disease, patients perceived a negative impact on their personal life, family, work, and community., Conclusions: Lack of knowledge regarding tuberculosis is an important factor in the selection of and adherence to different care alternatives. Inadequate care provided by health services, including an unsatisfactory physician-patient relationship, resulted in diagnostic delay and non-adherence to therapy. Education programs to promote basic knowledge regarding tuberculosis and its treatment are necessary in this region.

Objective: To estimate the effect of contraception on fertility in the border region of Chiapas, Mexico., Materials and Methods: In 1994 an epidemiological cross-sectional study was carried out on a representative sample of 1,560 non-indigenous women between ages 15 and 49 in the border region of Chiapas. The prevalence of contraception practices and the total fertility rates (TFR) were obtained and stratified by rural, intermediate and urban communities. TFR were compared between women who had never used contraceptives and those who had used them., Results: The estimated TFR was 3.67 and varied from 4.14 in rural areas to 3.36 in urban areas. There were no differences in the TFR (3.74 and 3.88) nor in the average live births (3.47 and 3.48) between women who had never used contraceptives and those who had used them., Conclusions: The major effect of contraception on fertility was observed in rural areas. Factors which influence the small impact of contraception on fertility include the late use of these methods and the early age of first union among users.

Objectives: To determine risk factors for antibiotic resistance in patients with pulmonary tuberculosis in four Health Jurisdictions of the state of Chiapas., Material and Methods: A case-control study was carried out in patients diagnosed by acid fast smear during 1992. A questionnaire was applied which included variables related to the diagnosis, treatment and follow-up of the patients. Sputum samples were collected for culture and sensitivity tests. A case of drug-resistant pulmonary tuberculosis was defined as the presence of culture colonies showing resistance to one or more drugs. The control group was patients with negative smears and cultures or positive cultures for M. tuberculosis sensitive to the specific drugs., Results: Of the total of 18 individuals reported to have positive cultures, 13 (72.2%) were resistant to one or more drugs. Resistance to two or more drugs was found in 10 of them of which three were resistant to five antituberculosis drugs. The most frequent resistance was to isoniazid (77%). Risk factors for resistance encountered in this patient population were monotherapy (OR = 34.2), abandonment of treatment (OR = 6.86), a prolonged period of illness (OR = 6.40), delay in diagnosis and a history of prior therapy (OR = 28.3)., Conclusions: The high proportion of patients resistant to antituberculosis therapy poses a public health problem and is a clear consequence of the problems arising from inadequate treatment.

Despite the widespread use of Bin 19 as a vector for plant transformation, detailed sequence information on its T-DNA region has only recently become available. We now show that the non-T-DNA region, like the T-DNA region, contains several superfluous insertions and find that some functional elements may not contain optimal sequences. Knowledge of the complete 11,777 bp sequence will aid in the construction of exceptionally efficient derivative vectors of approximately half this size. Precise knowledge of restriction sites and removal of unnecessary sequences will facilitate plasmid manipulations and plant transformation.

Much research in psychology has evaluated the quality of people's decisions by comparisons with subjective expected utility (SEU) theory. This article suggests that typical arguments made for the status of utility theory as normative do not justify its use by psychologists as a standard by which to evaluate decision quality. It is argued that to evaluate decision quality, researchers need to identify those decision processes that tend to lead to desirable outcomes. It is contended that a good decision-making process must be concerned with how (and whether) decision makers evaluate potential consequences of decisions, the extent to which they accurately identify all relevant consequences, and the way in which they make final choices. Research that bears on these issues is reviewed.

Dihydrodipicolinate synthase (DHPS; EC 4.2.1.52) is the first committed enzyme in the lysine branch of the aspartate-derived amino acid biosynthesis pathway and is common to bacteria and plants. Due to feedback inhibition by lysine, DHPS serves in a regulatory role for this pathway in plant metabolism. To elucidate the molecular genetic characteristics of DHPS, we isolated a putative full-length cDNA clone for maize DHPS by direct genetic selection in an Escherichia coli dapA- auxotroph. The maize DHPS activity expressed in the complemented E. coli auxotroph showed the lysine inhibition characteristics of purified maize DHPS, indicating that the cDNA encoded sequences for both the catalytic function and regulatory properties of the enzyme. The N-terminal amino acid sequence of purified maize DHPS was determined by direct sequencing and showed homology to a sequence within the cDNA, indicating that the clone contained the entire coding region for a mature polypeptide of 326 amino acids plus a 54 amino acid transit peptide sequence. The molecular weight of 35,854, predicted from the deduced amino acid sequence, was similar to the 38,000 Mr determined by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) for the purified enzyme from maize. DHPS mRNAs complementary to the cDNA were detected in RNA isolated from developing maize endosperm and embryo tissues. Southern blots indicated the presence of more than one genomic sequence homologous to DHPS per haploid maize genome.

Dihydrodipicolinate synthase (EC 4.2.1.52), the first enzyme specific to lysine biosynthesis in plants, was purified from maize (Zea mays L.) cell suspension cultures and leaves. The subunit molecular weight of maize dihydrodipicolinate synthase was estimated to be 38,000 based on SDS-PAGE. The condensation of l-aspartate semialdehyde and pyruvate by highly purified dihydrodipicolinate synthase exhibited kinetics characteristic of a Ping Pong Bi Bi ordered reaction in which pyruvate binds first to the enzyme. Substrate inhibition evident at higher concentrations of l-aspartate semialdehyde was partially alleviated by increasing concentrations of pyruvate. Pyruvate binding exhibited cooperativity with an apparent number of 2 and 1.86 millimolar concentration required for 50% of maximal activity. The K(m) for aspartate semialdehyde was estimated to be 0.6 millimolar concentration. Lysine was an allosteric cooperative inhibitor of dihydrodipicolinate synthase with an estimated Hill number of 4 and 23 micromolar concentration required for 50% inhibition. The physical and kinetic data are consistent with a homotetramer model for the native enzyme.

Aspartate kinase (AK; EC 2.7.2.A) catalyzes the first reaction in the biosynthesis pathway for aspartate-derived amino acids in plants. Aspartate kinase was purified from wildtype and two maize (Zea mays L.) genotypes carrying unlinked dominant mutations,Ask LT19 andAsk2 -LT20, that conferred overproduction of threonine, lysine, methionine and isoleucine. The objective of this investigation was to characterize the AKs from mutant and wildtype plants to determine their role in regulating the synthesis of aspartate-derived amino acids in maize. Kernels of the homozygousAsk2 mutant exhibited 174-, 10-, 13- and 2-fold increases in, in this sequence, free threonine, lysine, methionine and isoleucine, compared to wildtype. In wildtype maize, AK was allosterically feedback-inhibited by lysine with 10 μML-lysine required for 50% inhibition. In contrast, AK purified from the isogenic heterozygousAsk and homozygousAsk2 mutants required 25 and 760 μM lysine for 50% inhibition, respectively, indicating thatAsk andAsk2 were separate structural loci for lysine-regulated AK subunits in maize. Further characterization of purified AK from the homozygous mutantAsk2 line indicated altered substrate and lysine inhibition kinetics. The apparent Hill coefficient was 0.7 for the mutantAsk2 AK compared with 1.6 for the wildtype enzyme, indicating that the mutant allele conferred the loss of a lysinebinding site to the mutant AK. Lysine appeared to be a linear noncompetitive inhibitor ofAsk2 AK with respect to MgATP and an uncompetitive inhibitor with respect to aspartate compared to S-parabolic, I parabolic noncompetitive inhibition of wildtype AK. Reduced lysine sensitivity of theAsk2 gene product appeared to reduce the lysine inhibition of all of the AK activity detected in homozygousAsk2 plants, indicating that maize AK is a heteromeric enzyme consisting of the two lysine-sensitive polypeptides derived from theAsk andAsk2 structural genes.

1. An analytical procedure has been developed for the gas-liquid chromatographic analyses of cholesterol in hair samples. 2. The mean and standard deviation of hair samples from 40 different dogs have been established to be 632 +/- 291 ppm. No significant difference in cholesterol levels was observed between dogs of different breeds, sex or age.

Pigmented villonodular tenosynovitis is a proliferative disorder of the synovium that can involve the joints, tendon sheaths, and bursae. There are two histologically similar lesions of pigmented villonodular tenosynovitis--nodular pigmented villonodular tenosynovitis and diffuse pigmented villonodular tenosynovitis. The authors present a case involving nodular pigmented villonodular tenosynovitis, which is the more rare form of these two lesions.