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The annelid genus Diopatra occurs in all major oceans but is best represented in the shallow depths of warmer waters, where it lives in elaborately decorated tubes. This paper provides an introduction to the animals, discussing their history and diversity. We describe and illustrate its morphology and geographic distribution. While they were thought to be predominantly gonochoristic, recent reproductive studies show that several species are protandric simultaneous hermaphrodites. Development is by broadcast spawning with a brief pelagic stage or direct development in the parental tube or egg mass attached to it. Diopatra is a key ecosystem engineer, altering water flow and deposition and increasing the availability of refugia. We also discuss its harvesting as fishing bait, its role as an alien or introduced species, its capacity to regenerate, its therapeutic potential, and its applications as a bioindicator species for climate change, geographic distribution changes, and dispersal.

Opportunistic infections by Burkholderia cenocepacia are life threatening for patients suffering from cystic fibrosis and chronic granulomatous disease. These infections are often associated with variable clinical outcomes, prompting an interest in molecular investigations of phenotypes associated with disease severity. The production of the pyomelanin pigment is one such phenotype, which was recently linked to the ability of clinical strains to carry out biotransformation of the antibiotic trimethoprim. However, this biotransformation product was not identified, and differences in metabolite production associated with pyomelanin pigmentation are poorly understood. Here, we identify several key metabolites produced exclusively by the pyomelanin-producing strains. To provide insight into the structures and biosynthetic origin of these metabolites, we developed a mass spectrometry-based strategy coupling unsupervised in silico substructure prediction with stable isotope labeling referred to as MAS-SILAC (Metabolite Annotation assisted by Substructure discovery and Stable Isotope Labeling by Amino acids in Cell culture). This approach led to discovery of homogentisic acid as a precursor for biosynthesis of several natural products and for biotransformation of trimethoprim, representing a previously unknown mechanism of antibiotic tolerance. This work presents application of computational methods for analysis of untargeted metabolomic data to link the chemotype of pathogenic microorganisms with a specific phenotype. The observations made in this study provide insights into the clinical significance of the melanated phenotype.

Ligand-induced endocytosis of the immune receptor FLAGELLIN SENSING2 (FLS2) is critical for maintaining its proper abundance in the plasma membrane (PM) to initiate and subsequently down regulate cellular immune responses to bacterial flagellin or flg22-peptide. The molecular components governing PM abundance of FLS2, however, remain mostly unknown. Here, we identified Arabidopsis (Arabidopsis thaliana) DYNAMIN-RELATED PROTEIN1A (DRP1A), a member of a plant-specific family of large dynamin GTPases, as a critical contributor to ligand-induced endocytosis of FLS2 and its physiological roles in flg22-signaling and immunity against Pseudomonas syringae pv. tomato DC3000 bacteria in leaves. Notably, drp1a single mutants displayed similar flg22-defects as those previously reported for mutants in another dynamin-related protein, DRP2B, that was previously shown to colocalize with DRP1A. Our study also uncovered synergistic roles of DRP1A and DRP2B in plant growth and development as drp1a drp2b double mutants exhibited severely stunted roots and cotyledons, as well as defective cell shape, cytokinesis, and seedling lethality. Furthermore, drp1a drp2b double mutants hyperaccumulated FLS2 in the PM prior to flg22-treatment and exhibited a block in ligand-induced endocytosis of FLS2, indicating combinatorial roles for DRP1A and DRP1B in governing PM abundance of FLS2. However, the increased steady-state PM accumulation of FLS2 in drp1a drp2b double mutants did not result in increased flg22 responses. We propose that DRP1A and DRP2B are important for the regulation of PM-associated levels of FLS2 necessary to attain signaling competency to initiate distinct flg22 responses, potentially through modulating the lipid environment in defined PM domains., (© American Society of Plant Biologists 2021. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

In Australia, the deep-water (bathyal and abyssal) benthic invertebrate fauna is poorly known in comparison with that of shallow (subtidal and shelf) habitats. Benthic fauna from the deep eastern Australian margin was sampled systematically for the first time during 2017 RV 'Investigator' voyage 'Sampling the Abyss'. Box core, Brenke sledge, and beam trawl samples were collected at one-degree intervals from Tasmania, 42°S, to southern Queensland, 24°S, from 900 to 4800 m depth. Annelids collected were identified by taxonomic experts on individual families around the world. A complete list of all identified species is presented, accompanied with brief morphological diagnoses, taxonomic remarks, and colour images. A total of more than 6000 annelid specimens consisting of 50 families (47 Polychaeta, one Echiura, two Sipuncula) and 214 species were recovered. Twenty-seven species were given valid names, 45 were assigned the qualifier cf., 87 the qualifier sp., and 55 species were considered new to science. Geographical ranges of 16 morphospecies extended along the eastern Australian margin to the Great Australian Bight, South Australia; however, these ranges need to be confirmed with genetic data. This work providing critical baseline biodiversity data on an important group of benthic invertebrates from a virtually unknown region of the world's ocean will act as a springboard for future taxonomic and biogeographic studies in the area., (Laetitia M. Gunton, Elena K. Kupriyanova, Tom Alvestad, Lynda Avery, James A. Blake, Olga Biriukova, Markus Böggemann, Polina Borisova, Nataliya Budaeva, Ingo Burghardt, Maria Capa, Magdalena N. Georgieva, Christopher J. Glasby, Pan-Wen Hsueh, Pat Hutchings, Naoto Jimi, Jon A. Kongsrud, Joachim Langeneck, Karin Meißner, Anna Murray, Mark Nikolic, Hannelore Paxton, Dino Ramos, Anja Schulze, Robert Sobczyk, Charlotte Watson, Helena Wiklund, Robin S. Wilson, Anna Zhadan, Jinghuai Zhang.)

The Burkholderia cepacia complex is a group of closely related bacterial species with large genomes that infect immunocompromised individuals and those living with cystic fibrosis. Some of these species are found more frequently and cause more severe disease than others, yet metabolomic differences between these have not been described. Furthermore, our understanding of how these species respond to antibiotics is limited. We investigated the metabolomics differences between three most prevalent Burkholderia spp. associated with cystic fibrosis: B. cenocepacia , B. multivorans , and B. dolosa in the presence and absence of the antibiotic trimethoprim. Using a combination of supervised and unsupervised metabolomics data visualization and analysis tools, we describe the overall differences between strains of the same species and between species. Specifically, we report, for the first time, the role of the pyomelanin pathway in the metabolism of trimethoprim. We also report differences in the detection of known secondary metabolites such as fragin, ornibactin, and N -acylhomoserine lactones and their analogs in closely related strains. Furthermore, we highlight the potential for the discovery of new secondary metabolites in clinical strains of Burkholderia spp. The metabolomics differences described in this study highlight the personalized nature of closely related Burkholderia strains.

Two new species of Mooreonuphis, a genus best known from the coasts of the Americas, are described from Australia, bringing the number of recognized Mooreonuphis species to 23. Mooreonuphis ariasi n. sp. was collected from off Sydney to Wollongong, New South Wales in sandy sediments, in depths of 25-50 m and M. wilsoni n. sp. in the Bass Strait and Tasmania, in 89-130 m. The two new Australian species are among the smallest and the only known abranchiate species in the genus. The summer collections of M. ariasi n. sp. contained a large number of brooders with their young undergoing asynchronous direct development in the parental tube. Nothing is known about the development of M. wilsoni n. sp.

Background: Recent evidence suggests that autonomic nervous dysfunction may be one of many potential factors contributing to persisting post-concussion symptoms., Objective: This is the first systematic review to explore the impact of concussion on multiple aspects of autonomic nervous system functioning., Methods: The methods employed are in compliance with the American Academy of Neurology (AAN) and PRISMA standards. Embase, MEDLINE, PsychINFO, and Science Citation Index literature searches were performed using relevant indexing terms for articles published prior to the end of December 2016. Data extraction was performed by two independent groups, including study quality indicators to determine potential risk for bias according to the 4-tiered classification scheme of the AAN., Results: Thirty-six articles qualified for inclusion in the analysis. Only three studies (one Class II and two Class IV) did not identify anomalies in measures of ANS functioning in concussed populations., Conclusions: The evidence supports the conclusion that it is likely that concussion causes autonomic nervous system anomalies. An awareness of this relationship increases our understanding of the physical impact of concussion, partially explains the overlap of concussion symptoms with other medical conditions, presents opportunities for further research, and has the potential to powerfully inform treatment decisions.

The genus Aponuphis, previously known from the eastern North Atlantic and the Mediterranean, is newly reported from Australia. Three new species are described from off Sydney to Wollongong, New South Wales in sandy sediments, in depths of 25-70 m, bringing the number of recognized Aponuphis species to ten. Aponuphis annae n. sp. and A. bellani n. sp. are abranchiate, whilst A. danicae n. sp. has branchiae over a short region of its body. The tubes of A. annae and A. danicae have a fragile mucous consistency with attached sand grains and that of A. bellani is transparent, tight-fitting and smooth. Two specimens of A. annae were collected with developing juveniles in their tubes demonstrating their direct development but the reproductive mode of the other two species is not known. The distinguishing characteristics of all recognized species are tabled and a key to the three Australian species is presented.

Examination of the holotype of Paradiopatra bihanica (Intes & LeLoeuff, 1975) indicated that the identity of this species had been incorrectly represented in recent papers and that the ensuing synonymisation of P. calliopae Arvanitidis & Koukouras, 1997 was unwarranted. This led us to researching the history of the holotype and its representation in the literature to reinstate P. calliopae as the correct name for this very common Mediterranean and eastern North Atlantic species (Martínez & Adarraga 2001; Arias & Paxton 2015; Santelli et al. 2015).

This work recognises five species of Paradiopatra from southwestern Europe, represented in the Bay of Biscay, the Atlantic coasts of Iberia and the western Mediterranean Sea. One species, Paradiopatra florencioi sp. nov., is newly described from the Cantabrian slope, northern Spain. We are presenting detailed diagnoses of all species and report ontogenetic changes of P. bihanica and P. hispanica. Scanning electron microscopy elucidated previously known Methylene Blue or Green staining patterns of the ventral glandular pads as cuticular pores. A dichotomous key to all species is included.

This study describes a new subgenus of miniature, progenetic Rhamphobrachium species from eastern Australia. Minibrachium, n. subg. is characterised by such paedomorphic features as lack of peristomial cirri and branchiae, possession of only two pairs of modified parapodia with spiny recurved hooks and very early onset of subacicular hooks. Three new species are described, of which at least R. (Minibrachium) nutrix n. sp., the type species (from Lizard Island, Great Barrier Reef), is a fully grown adult, brooding several 16- to 20-chaetiger juveniles in the parental tubes. Rhamphobrachium (M.) talboti n. sp. (from off Sydney) and R. (M.) fractum n. sp. (from Bass Strait), although twice as large as the type species, have only partially developed frontal lips, indicating either incomplete growth or presenting an additional paedomorphic character. We discuss the position of the new subgenus in the Rhamphobrachium complex and present a key to the subgenera of Rhamphobrachium and species of R. (Minibrachium).

This study recognises five species of Onuphis and four species of Aponuphis from southwestern Europe, represented in the Bay of Biscay, the Atlantic coasts of Iberia and the western Mediterranean Sea. One species, Onuphis anadonae sp. nov., is newly described and another species, Aponuphis willsiei Cantone & Bellan, 1996 is a new record for the Iberian Peninsula waters. Aponuphis ornata (Fauvel, 1928) and A. willsiei are shown to be tube brooders and the development of the juveniles is described. The usefulness of some Onuphis characters, as well as the size-dependent distribution of branchiae and chaetae, complicating Aponuphis species identification, are discussed. We are presenting descriptions and photographs of the distinctive Aponuphis colour pattern which we recommend as an identificatory tool instead. A dichotomous key to all species is included.

Onuphis eremita, type species of the genus, and Onuphis pancerii, originally from La Rochelle (France) and Gulf of Naples (Italy) respectively, are two of the earliest described species of the family Onuphidae. Yet, the definition of the former was extremely confused, the latter was considered its synonym for nearly a century, and types do not exist. We have examined a large amount of material, old museum collections and newly collected specimens from the European Atlantic and Mediterranean and conclude that both, O. eremita and O. pancerii, are valid species. The most obvious differences are that O. eremita is a slender species with only tridentate pseudocompound hooks while O. pancerii is much more robust and has pseudocompound hooks with bi- and tridentate (or even multidentate) tips. Neotypes have been designated for both species and the two taxa are redescribed and illustrated. We regard O. eremita sensu stricto, as redefined on the neotype here, to be restricted to the European Atlantic and western and central Mediterranean; other records need to be re-evaluated. Some specimens of both species were collected with eggs (~ 200 µm diameter) protruding through openings in the body wall, and some O. eremita had ect-aquasperm, suggesting that reproduction is by broadcast spawning and indirect development.

In broiler chickens, genetic success for desired production traits is often shadowed by welfare concerns related to musculoskeletal health. Whilst these concerns are clear, a viable solution is still elusive. Part of the solution lies in knowing how anatomical changes in afflicted body systems that occur across ontogeny influence standing and moving. Here, to demonstrate these changes we quantify the segment inertial properties of the whole body, trunk (legs removed) and the right pelvic limb segments of five broilers at three different age groups across development. We also consider how muscle architecture (mass, fascicle length and other properties related to mechanics) changes for selected muscles of the pelvic limb. All broilers used had no observed lameness, but we document the limb pathologies identified post mortem, since these two factors do not always correlate, as shown here. The most common leg disorders, including bacterial chondronecrosis with osteomyelitis and rotational and angular deformities of the lower limb, were observed in chickens at all developmental stages. Whole limb morphology is not uniform relative to body size, with broilers obtaining large thighs and feet between four and six weeks of age. This implies that the energetic cost of swinging the limbs is markedly increased across this growth period, perhaps contributing to reduced activity levels. Hindlimb bone length does not change during this period, which may be advantageous for increased stability despite the increased energetic costs. Increased pectoral muscle growth appears to move the centre of mass cranio-dorsally in the last two weeks of growth. This has direct consequences for locomotion (potentially greater limb muscle stresses during standing and moving). Our study is the first to measure these changes in the musculoskeletal system across growth in chickens, and reveals how artificially selected changes of the morphology of the pectoral apparatus may cause deficits in locomotion.

Genetic selection for improved meat yields, digestive efficiency and growth rates have transformed the biology of broiler chickens. Modern birds undergo a 50-fold multiplication in body mass in just six weeks, from hatching to slaughter weight. However, this selection for rapid growth and improvements in broiler productivity is also widely thought to be associated with increased welfare problems as many birds suffer from leg, circulatory and respiratory diseases. To understand growth-related changes in musculoskeletal and organ morphology and respiratory skeletal development over the standard six-week rearing period, we present data from post-hatch cadaveric commercial broiler chickens aged 0, 2, 4 and 6 weeks. The heart, lungs and intestines decreased in size for hatch to slaughter weight when considered as a proportion of body mass. Proportional liver size increased in the two weeks after hatch but decreased between 2 and 6 weeks. Breast muscle mass on the other hand displayed strong positive allometry, increasing in mass faster than the increase in body mass. Contrastingly, less rapid isometric growth was found in the external oblique muscle, a major respiratory muscle that moves the sternum dorsally during expiration. Considered together with the relatively slow ossification of elements of the respiratory skeleton, it seems that rapid growth of the breast muscles might compromise the efficacy of the respiratory apparatus. Furthermore, the relative reduction in size of the major organs indicates that selective breeding in meat-producing birds has unintended consequences that may bias these birds toward compromised welfare and could limit further improvements in meat-production and feed efficiency.

Diagnosis and management of traumatic brain injury (TBI) is crucial to improve patient outcomes. While initial head computed tomography (CT) scan is the optimum tool for quick and accurate detection of intracranial hemorrhage, the guidelines on use of repeat CT differ among institutions. Three systematic reviews have been conducted on a similar topic; none have performed a comprehensive meta-analysis of all studies. Search of Medline, the Cochrane Library database, and Clinicaltrials.gov , and a hand search of conference abstracts and references for all completed studies reporting data on change in management following repeat CT was conducted. Two authors reviewed all studies and extracted data using a standardized form. A proportional meta-analysis was conducted using the random-effects model for outcomes related to any change in management following repeat CT. Any change in management included intracranial intervention, change in intracranial pressure monitoring, and/or administration of drug therapy. Search results yielded 6982 references. In all, 41 studies enrolling 10,501 patients were included. Change in management following repeat CT was reported in 13 prospective and 28 retrospective studies and yielded a pooled proportion of 11.4% (95% confidence interval [CI] 5.9-18.4) and 9.6% (95% CI 6.5-13.2), respectively. In a subgroup analysis of mild TBI patients (Glasgow Coma Scale score 13 to 15), five prospective and nine retrospective studies reported on change in management following repeat CT with the pooled proportion across prospective studies at 2.3% (95% CI 0.3-6.3) and across retrospective studies at 3.9% (95% CI 2.3-5.7), respectively. The evidence suggests that repeat CT in patients with TBI results in a change in management for only a minority of patients. Better designed studies are needed to address the issue of the value of repeat CT in the management of TBI.

A new species of the genus Mooreonuphis Fauchald, 1982 collected from the Cantabrian shelf (Bay of Biscay) is described. Mooreonuphis vespa sp. nov. constitutes the first record of this genus in European waters and is characterised by: conspicuous dark brown transverse segmental pigment bands; antennae and palps with 3-5 basal ceratophoral rings and a very long distal ring; unusually long and slender peristomial and dorsal cirri; simple branchiae from chaetiger 17-19. We present observations on its reproductive biology (including brooding behaviour) and remark on the biogeography of the genus.

One of the most extraordinary results of selective breeding is the modern broiler chicken, whose phenotypic attributes reflect its genetic success. Unfortunately, leg health issues and poor walking ability are prevalent in the broiler population, with the exact aetiopathogenesis unknown. Here we present a biomechanical analysis of the gait dynamics of the modern broiler and its two pureline commercial broiler breeder lines (A and B) in order to clarify how changes in basic morphology are associated with the way these chickens walk. We collected force plate and kinematic data from 25 chickens (market age), over a range of walking speeds, to quantify the three-dimensional dynamics of the centre of mass (CoM) and determine how these birds modulate the force and mechanical work of locomotion. Common features of their gait include extremely slow walking speeds, a wide base of support and large lateral motions of the CoM, which primarily reflect changes to cope with their apparent instability and large body mass. These features allowed the chickens to keep their peak vertical forces low, but resulted in high mediolateral forces, which exceeded fore-aft forces. Gait differences directly related to morphological characteristics also exist. This was particularly evident in Pureline B birds, which have a more crouched limb posture. Mechanical costs of transport were still similar across all lines and were not exceptional when compared with more wild-type ground-running birds. Broiler chickens seem to have an awkward gait, but some aspects of their dynamics show rather surprising similarities to other avian bipeds.

Background: The Emergency Department Algorithm (EDA) developed at New York University uses administrative discharge data to distill hundreds of International Classification of Diseases-9 codes for emergency department (ED) visits into 4 categories, making it attractive to researchers and policy makers. The EDA has been used to analyze patterns of ED visits in a wide variety of locations and populations. However, there are concerns regarding the validity and use of the EDA for research and policy., Objective: To explain the findings of previous EDA users that it appears to lack sensitivity in detecting changes in ED utilization patterns., Study Design: Mathematical simulation was used to analyze and explain the performance of the EDA in detecting differences in utilization patterns across hypothetical ED populations. Sensitivity analysis was used to illustrate the magnitude of changes in EDA outputs relative to changes in ED populations using a national sample of actual ED patients., Results: The vast majority of possible EDA outputs are clustered so tightly as to show no significant change in outputs between different hypothetical populations. Sensitivity analysis shows that changes in EDA outputs are not nearly as great as the magnitude of the input differences across real-world populations., Conclusions: The EDA categorizes a very large variety of ED visits into a relatively small group of outputs. Its operating characteristics suggest that the EDA is insufficiently sensitive to changes in ED utilization patterns to be useful in assessing interventions to change them. This finding should caution potential users to consider the EDA's limitations before using it.

Background: There is a growing need for palliative care services located outside of hospitals., Objective: This study's objective was to evaluate a home-based, nonhospice, palliative medicine (PM) consultation practice within a fee-for-service environment., Method: Hospital and emergency department (ED) utilization and cost data obtained from administrative records were analyzed with longitudinal analyses to compare use 18 months before and after service enrollment in a single patient group., Participants: Patients (N=369) with advanced complex illness (ACI) referred for home-based palliative consultation participated in the study., Intervention: Consultation conducted by nurse practitioners included a multidimensional assessment with recommendations to outpatient physicians for symptom management and guidance to patient and family for goals of treatment and advanced care planning (ACP). Nurse practitioners were supported by a collaborating PM physician. Follow-up visits varied by need for symptom management and ACP., Results: Total hospitalizations, total hospital days, total and variable costs, and probability of a 30-day readmission were significantly reduced in the 18-month period following program enrollment. However, probability of an ED visit was not reduced., Conclusions: While requiring replication with rigorous methods, preliminary results suggest a home-based PM practice may reduce hospital utilization for ACI patients.

This study recognises eight species of Paradiopatra from eastern Australian waters, of which only one, Paradiopatra ehlersi, had been previously recorded. Six species are newly described: P. acirrata n. sp., P. imajimai n. sp., P. longicappa n. sp., P. piccola n. sp., P. spinosa n. sp., P. variabilis n. sp., and one species is reported as P. cf. papillata. Descriptions of all species (with the exception of P. ehlersi) are given, accompanied by detailed illustrations, including scanning electron micrographs and drawings. A key to all 32 species of the genus is included plus a table summarising specific diagnostic characters of the Australian representatives.

Introduction of an evidence-based practice change, such as hourly rounding, can be difficult in the hospital setting. This study used ethnographic methods to examine problems with the implementation of hourly rounding on 2 similar inpatient units at our hospital. Results indicate that careful planning, communication, implementation, and evaluation are required for successful implementation of a nursing practice change.

Intensive artificial selection has led to the production of the modern broiler chicken, which over the last few decades has undergone a dramatic increase in growth rate and noticeable changes in body conformation. Unfortunately, this has been associated with musculoskeletal abnormalities which have altered the walking ability of these birds, raising obvious welfare concerns, as well as causing economic losses. Here we present a comparative study of ancestral and derived muscle anatomy in chickens to begin to tease apart how evolutionary alterations of muscle form in chickens have influenced their locomotor function and perhaps contributed to lameness. We measured the muscle architectural properties of the right pelvic limb in 50 birds, including the Giant Junglefowl, a commercial strain broiler and four pureline commercial broiler breeder lines (from which the broiler populations are derived) to identify which features of the broiler's architectural design have diverged the most from the ancestral condition. We report a decline in pelvic limb muscle mass in the commercial line birds that may compromise their locomotor abilities because they carry a larger body mass. This greater demand on the pelvic limb muscles has mostly led to changes in support at the hip joint, revealing significantly larger abductors and additionally much larger medial rotators in the broiler population. Differences were seen within the commercial line bird populations, which are likely attributed to different selection pressures and may reflect differences in the walking ability of these birds. In addition, Junglefowl seem to have both greater force-generating capabilities and longer, presumably faster contracting muscles, indicative of superior musculoskeletal/locomotor function. We have provided baseline data for generating hypotheses to investigate in greater depth the specific biomechanical constraints that compromise the modern broiler's walking ability and propose that these factors should be considered in the selection for musculoskeletal health in the chickens of the future. Our new anatomical data for a wide range of domestic and wild-type chickens is useful in a comparative context and for deeper functional analysis including computer modelling/simulation of limb mechanics.

Background: Little evidence exists to support implementing various health information technologies, such as telemedicine, in intensive care units., Methods: A coordinated health information technology bundle (HITB) was implemented along with remote intensivist coverage (RIC) at a 727-bed academic community hospital. Critical care specialists provided bedside coverage during the day and RIC at night to achieve intensivist coverage 24 hours per day, 7 days per week. We evaluated the effect of HITB-RIC on mortality, ventilator and vasopressor use, and the intervention length of stay. We compared our results with those achieved at baseline., Results: A total of 954 control patients who received care for 16 months before the implementation of HITB-RIC and 959 study patients who received care for 10 months after the implementation were included in the analysis. Mortality for the control and intervention groups were 21.4% and 14.7%, respectively. In addition, the observed mortality for the intervention group was 75.8% (P < .001) of that predicted by the Acute Physiology and Chronic Health Evaluation IV hospital mortality equations, which was 29.5% lower relative to the control group. Regression results confirm that the hospital mortality of the intensive care unit patients was significantly lower after implementation of the intervention, controlling for predicted risk of mortality and do-not-resuscitate status. Overall, intervention patients also had significantly less (P = .001) use of mechanical ventilation, controlling for body-system diagnosis category and severity of illness., Conclusion: The use of HITB-RIC was associated with significantly lower mortality and less ventilator use in critically ill patients.

Inertial properties of animal bodies and segments are critical input parameters for biomechanical analysis of standing and moving, and thus are important for paleobiological inquiries into the broader behaviors, ecology and evolution of extinct taxa such as dinosaurs. But how accurately can these be estimated? Computational modeling was used to estimate the inertial properties including mass, density, and center of mass (COM) for extant crocodiles (adult and juvenile Crocodylus johnstoni) and birds (Gallus gallus; junglefowl and broiler chickens), to identify the chief sources of variation and methodological errors, and their significance. High-resolution computed tomography scans were segmented into 3D objects and imported into inertial property estimation software that allowed for the examination of variable body segment densities (e.g., air spaces such as lungs, and deformable body outlines). Considerable biological variation of inertial properties was found within groups due to ontogenetic changes as well as evolutionary changes between chicken groups. COM positions shift in variable directions during ontogeny in different groups. Our method was repeatable and the resolution was sufficient for accurate estimations of mass and density in particular. However, we also found considerable potential methodological errors for COM related to (1) assumed body segment orientation, (2) what frames of reference are used to normalize COM for size-independent comparisons among animals, and (3) assumptions about tail shape. Methods and assumptions are suggested to minimize these errors in the future and thereby improve estimation of inertial properties for extant and extinct animals. In the best cases, 10%-15% errors in these estimates are unavoidable, but particularly for extinct taxa errors closer to 50% should be expected, and therefore, cautiously investigated. Nonetheless in the best cases these methods allow rigorous estimation of inertial properties., ((c) 2009 Wiley-Liss, Inc.)

As the largest extant terrestrial animals, elephants do not trot or gallop but can move smoothly to faster speeds without markedly changing their kinematics, yet with a shift from vaulting to bouncing kinetics. To understand this unusual mechanism, we quantified the forelimb and hindlimb motions of eight Asian elephants (Elephas maximus) and seven African elephants (Loxodonta africana). We used 240 Hz motion analysis (tracking 10 joint markers) to measure the flexion/extension angles and angular velocities of the limb segments and joints for 288 strides across an eightfold range of speeds (0.6-4.9 m s(-1)) and a sevenfold range of body mass (521-3684 kg). We show that the columnar limb orientation that elephants supposedly exemplify is an oversimplification--few segments or joints are extremely vertical during weight support (especially at faster speeds), and joint flexion during the swing phase is considerable. The 'inflexible' ankle is shown to have potentially spring-like motion, unlike the highly flexible wrist, which ironically is more static during support. Elephants use approximately 31-77% of their maximal joint ranges of motion during rapid locomotion, with this fraction increasing distally in the limbs, a trend observed in some other running animals. All angular velocities decrease with increasing size, whereas smaller elephant limbs are not markedly more flexed than adults. We find no major quantitative differences between African and Asian elephant locomotion but show that elephant limb motions are more similar to those of smaller animals, including humans and horses, than commonly recognized. Such similarities have been obscured by the reliance on the term ;columnar' to differentiate elephant limb posture from that of other animals. Our database will be helpful for identifying elephants with unusual limb movements, facilitating early recognition of musculoskeletal pathology.

Jaw shedding and replacement of Diopatra aciculata (Onuphidae, Eunicida), by the same process as arthropod molting, involving apolysis and ecdysis, is described here. These observations suggest that molting has either evolved convergently in eunicidan polychaetes and ecdysozoans or it was present in the last ecdysozoan/lophotrochozoan common ancestor and thus may not represent a synapomorphy of the ecdysozoans.

This study of a subset of women and infants participating in National Institutes of Health Pediatric AIDS Clinical Trials Group protocol 185 evaluated lymphocyte phenotypic markers of immune activation and differentiation to determine their association with the likelihood of human immunodeficiency virus (HIV) transmission from the women to their infants and the potential for early identification and/or prognosis of infection in the infants. Lymphocytes from 215 human immunodeficiency virus type 1 (HIV)-infected women and 192 of their infants were analyzed by flow cytometry with an extended three-color panel of monoclonal antibodies. Women who did not transmit to their infants tended to have higher CD4+ T cells. Most notably, levels of total CD8+ T cells and CD8+ CD38+ cells made significant independent contributions to predicting the risk of mother-to-child transmission. Adjusting for HIV-1 RNA level at entry, a one percentage-point increase in these marker combinations was associated with a nine percent increase in the likelihood of maternal transmission. Total as well as naive CD4+ T cells were significantly higher in uninfected than infected infants. Total CD8+ cells, as well as CD8+ cells positive for HLA-DR+, CD45 RA+ HLA-DR+, and CD28+ HLA-DR+ were elevated in infected infants. Detailed immunophenotyping may be helpful in predicting which pregnant HIV-infected women are at increased risk of transmitting HIV to their infants. Increasing differences in lymphocyte subsets between infected and uninfected infants became apparent as early as six weeks of age. Detailed immunophenotyping may be useful in supporting the diagnosis of HIV infection in infants with perinatal HIV exposure.

Background: The reported annual incidence of Rocky Mountain spotted fever in the United States is 2.2 per million, but studies have suggested that human infection with Rickettsia rickettsii may be more common. This study estimated the prevalence of antibodies reactive to R rickettsii among children living in the southeastern and south central United States., Study Design: Approximately 300 specimens were obtained from children at each of 7 pediatric referral centers (N = 1999). Serum was tested for R rickettsii antibodies by means of indirect immunofluorescence antibody assay. Three different cutoff titers (>or=64, >or=128, and >or=256) represented increasing levels of stringency to define positive specimens., Results: Overall, 12.0% of children had R rickettsii antibody titers of at least 64; 7.3%, at least 128; and 4.3%, at least 256. Strong relationships were seen between increasing age and seroprevalence at each cutoff titer. Remarkably, 6.4% of children aged 13 to 17 years had titers of at least 256. Age-adjusted seroprevalence rates at titers of at least 64 varied from 21.9% in Little Rock, Ark, to 3.5% in Louisville, Ky. At titers of at least 256, seroprevalence ranged from 7.7% in Nashville, Tenn, to 1.8% in Winston-Salem, NC. Only site and age group were strong predictors of seropositivity; a weak association was seen with nonurban residence., Conclusions: To our knowledge, this is the largest serosurvey of rickettsial infection in children in the United States. Within the limitations of the immunofluorescence antibody assay, these data suggest that infections with R rickettsii or antigenically related spotted-fever group rickettsiae may be common and subclinical. The results also have implications for the interpretation of single immunofluorescence antibody assay titers in children with suspected Rocky Mountain spotted fever.

Background: The reported annual incidence of human monocytic ehrlichiosis, which is due to infection with Ehrlichia chaffeensis, is as high as 5.5 per million in some states, but serosurveys suggest much higher infection rates in some populations., Objective: To estimate the prevalence of E chaffeensis infection among children aged 1 to 17 years living in the southeast and south-central United States., Design: Cross-sectional serosurvey., Setting: Seven academic pediatric medical centers in the southeastern and south-central United States., Patients: Nineteen hundred ninety-nine children (approximately 300 at each center) having their blood drawn for any reason., Main Outcome Measure: The presence of antibody at 2 different cutoff titers to E chaffeensis, as detected by indirect immunofluorescence assay., Results: Overall, 250 children (13%) had E chaffeensis antibody titers of 1:80 or higher and 61 (3%) had titers of 1:160 or higher. Age-adjusted seroprevalence rates varied widely between sites. At 1:80 or higher, the highest rate was in Winston-Salem, NC (22%), and the lowest was in Louisville, Ky (2%). At 1:160 or higher, the highest rate was in Kansas City, Mo (9%), and the lowest was in Oklahoma City, Okla (<1%). In univariate analyses, no associations were found between seroprevalence at either cutoff value and sex, race, source of specimen, or residence demographics. However, age was a significant predictor of seroprevalence at both cutoff values. In multiple logistic regression analysis, study site and age remained strong predictors of seroprevalence, but living in a nonurban ZIP code was not significantly related., Conclusion: Infection with E chaffeensis, or related ehrlichiae, may be more common in children than previously recognized.

Leptospirosis is a common and underdiagnosed zoonosis. Two rapid assays for serological diagnosis of acute leptospirosis in diagnostic laboratories, the immunoglobulin M (IgM)-dipstick assay and the indirect hemagglutination assay (IHA), were evaluated and compared with standard assays. Sera were examined from 104 patients admitted to a hospital for investigation in a leptospirosis diagnostic protocol. Specimens for serology were taken on days 1 and 4 of the patients' hospital stay. Antibodies were detected using an IgM-enzyme-linked immunosorbent assay (ELISA), microscopic agglutination test (MAT), an IgM-dipstick assay, and an IHA. Fifty-one patients were found to have leptospirosis. The sensitivity of the IgM-dipstick assay was 98%, its specificity was 90.6%, its positive predictive value was 90.9%, and its negative predictive value was 98%. The sensitivity of the IHA was 92.2%, its specificity was 94.4%, its positive predictive value was 95.9%, and its negative predictive value was 92.7%. The standard IgM-ELISA and MAT, were positive in the first samples tested from 67 and 55% of the cases, respectively, and the rapid IgM-dipstick assay and IHA were positive in 71 and 49%, respectively, in the first sample tested. Both rapid assays are highly sensitive and specific. Neither requires specialized equipment, and both are suitable for use in diagnostic laboratories.

Two easy-to-use commercial diagnostic assays, a dipstick enzyme-linked immunosorbent assay (ELISA) (Integrated Diagnostics, Baltimore, Md.) and an immunochromatographic card assay (PanBio, Brisbane, Australia) were evaluated for detection of immunoglobulin M (IgM) antibody to dengue virus with an in-house IgM antibody capture microplate ELISA as a reference assay. The dipstick ELISA was based on the indirect-ELISA format using dengue 2 virus as the only antigen and enzyme-labeled goat anti-human IgM antibody as the detector. The total assay time was 75 min. The immunochromatographic card assay was based on the antibody capture format and separately measured both anti-dengue virus IgM and IgG in the same test. Colloidal-gold-labeled anti-dengue virus monoclonal antibody bound with dengue virus 1 to 4 antigen cocktail was the detector, and anti-human IgM and IgG were the capture antibodies. The total assay time was <10 min. Sera from 164 individuals classified as either anti-dengue virus IgM positive (94) or anti-dengue virus IgM negative (70) in the reference microplate ELISA with a dengue virus 1 to 4 antigen cocktail were tested in the two commercial assays. The dipstick ELISA missed 7 of 94 positive samples, for a sensitivity of 92.6%, while the immunochromatographic card assay missed two positive samples, for a sensitivity of 97.9%. Of the 70 negative samples, four were false positive by the dipstick ELISA and two were false positive in the immunochromatographic card assay, resulting in specificities of 94.3 and 97.1%, respectively. Both commercial assays provide sensitive and specific detection of anti-dengue virus IgM antibody and could prove useful in settings where the microplate ELISA is impractical.

Purpose: To determine the prevalence of serologic reactivity, the 1-year incidence of seroconversion, and the frequency of multiple infections, and their associations with symptoms in a group of volunteers at high risk for tick-borne infections in New York state., Methods: We performed a seroepidemiologic study of Lyme borreliosis, 2 of the ehrlichioses, Rocky Mountain spotted fever, and babesiosis among 671 participants who lived or worked in a high-risk area (mainly in eastern Long Island, New York) for tick-borne diseases. Sera were collected in the winters of 1994 and 1995. Signs and symptoms of tick-borne disease were monitored monthly by mail and telephone. Lyme borreliosis serologies were done by enzyme-linked immunosorbent assay and Western blot. Rocky Mountain spotted fever serologies were initially screened using Dip-S-Ticks, followed by specific indirect immunofluorescence. Ehrlichiosis serologies were determined by epifluorescent microscopy, as were antibodies to Babesia microti., Results: Of the 671 participants, 88 (13%) had antibodies to > or = 1 tick-borne organisms, including 34 (5% of the total) with antibodies to Borrelia burgdorferi. Twenty-seven participants had evidence of exposure to B. burgdorferi at baseline. Seven participants (1%) seroconverted during the course of the study, 5 of whom were symptomatic for Lyme borreliosis. Antibodies to spotted fever group rickettsiae were seen in 28 participants (4%), 22 of whom were positive at baseline and 6 of whom seroconverted during the observation period. None of the seropositive patients had any symptoms or signs of infection. Twenty-four participants (3%) had serologic evidence of exposure to Ehrlichia (all but one to Ehrlichia equi); 5 (0.7%) seroconverted during the observation period, including 3 subjects who were asymptomatic. Antibodies to B. microti were seen in 7 participants (1%), including one asymptomatic seroconversion during the year of observation. There was evidence of possible dual infection in 5 patients., Conclusion: In a high-risk population, there was evidence of exposure to 5 tick-borne pathogens; however, many infections were asymptomatic, and coinfections were rare.

A rapid dipstick test for scrub typhus was prospectively evaluated in Chiangrai, northern Thailand. Sera from 162 patients with fever of unclear etiology were tested by a dot blot immunoassay using two different antigen concentrations. Dipsticks coated with lower concentration of antigen lacked sensitivity compared with the indirect immunoperoxidase test. Dipsticks with higher antigen concentration had increased sensitivity that was equivalent to that of the immunoperoxidase test. By increasing the antigen concentration on the dipstick, sensitivity increased from 67% to 100%, positive predictive value increased from 90% to 93%, and negative predictive value rose from 92% to 100%. The specificity of both antigen concentrations was 98%. This study establishes that scrub typhus can be confirmed serologically by use of a dipstick assay and that serodiagnosis can be effectively tailored to a target population.

The commercial dot-blot enzyme-linked immunosorbent assay Dip-S-Ticks dipstick test was compared with the indirect immunoperoxidase (IIP) and Weil-Felix (WF) tests for the diagnosis of scrub typhus, using the indirect immunofluorescent antibody test (IFA) as the reference standard. With a panel of 117 positive and 75 negative sera, the dipstick test was 94% sensitive and 98.7% specific at a cut-off value of one or more positive dots. The IIP was 90.6% sensitive and 100% specific at a cut-off titre of 1:400, and was more sensitive than the IFA with acute sera (79.6% vs. 68.5% at a titre > or = 1:400). All 3 were superior to the WF, which lacked sensitivity. The dipstick assay was easy to perform and did not require sophisticated electrical equipment, and the results were available within one hour. It is therefore suitable for use in rural Thailand, where scrub typhus is common.

Accurate serological confirmation of dengue (DEN) infection is difficult, because simple reliable assays for the detection of DEN antibodies are not available. To address this problem, a dipstick enzyme-linked immunosorbent assay (ELISA) was evaluated. The dipstick contained dots of serially diluted DEN 2 antigen. To detect immunoglobulin G (IgG), the dipstick was processed through four reaction cuvettes containing test serum, enhancer, enzyme-conjugated anti-human IgG and IgM antibody, and substrate. Total assay time was 45 min. To detect IgM, the serum was passed through a protein G device to remove IgG. The dipstick was then processed as before, except that the incubation times were longer and enzyme-conjugated anti-human IgM was used. The total assay time was 3 h. The dipstick ELISA results were compared with results from microplate ELISA. The IgG dipstick ELISA showed a sensitivity of 95.2% and a specificity of 100% compared to an IgG microplate ELISA with serum samples from 125 individuals living in an area in which DEN is endemic. In tests with 75 serum samples from patients with clinically suspected acute DEN infections, the IgM dipstick ELISA showed a sensitivity of 97.9% and specificity of 100% compared to those of an IgM antibody capture microplate ELISA. These results showed that the dipstick ELISA was a sensitive and specific test for the detection of either DEN IgM or IgG in human serum. The dipstick ELISA was also shown to be useful for detecting seroconversions to DEN IgM or IgG in paired serum samples from 20 patients with virus isolation-confirmed acute DEN infections.

We compared a commercially available dot-blot immunoassay system with the indirect immunofluorescence assay (IFA) in tests of known negative and known positive sera from scrub typhus cases. Using a panel of 100 sera from patients with various rickettsial and nonrickettsial infections, we observed that the IFA was 99% specific and the dipstick assay was 98% specific. In tests of 91 sera (30 negative and 61 positive for scrub typhus antibodies) from a study of febrile patients in Malaysia, using the standard of an IFA titer < 1:64 as negative, an IFA titer > 1:128 as positive, and an IFA titer = 1:64 as either positive or negative (supported by clinical records), dipsticks were 83% specific and 90% sensitive. The quantitative correlation of the dipsticks to IFA titers was confirmed by significant differences in geometric means of inverse IFA titers corresponding to the number of positive dipstick spots (no dots = 8.5, one dot = 43.3, two dots = 206.7, and three dots = 676.9). The assay would enable physicians and public health workers who deal with patients to quickly diagnose and appropriately treat most cases of the disease, especially in areas of high prevalence where the proportion of false-positive results to true-positive results would be low.

A commercial enzyme immunoassay kit called the Dip-S-Ticks (DS) for the detection of total immunoglobulin (Ig) G and IgM human antibodies to Rickettsia typhi was evaluated. In tests with 340 serum samples from patients with diagnosed cases of rickettsial diseases, patients suffering from other febrile illnesses, and normal subjects, the DS compared favorably with the standard indirect fluorescent-antibody (IFA) test. At IFA cutoff titers of > or = 1.64 and > or = 1:128, the DS showed sensitivities of 88.2 and 91.4% and specificities of 91.8 and 87.7%, respectively. The DS test correlated significantly with both the IFA IgG (r = 0.84, P < 0.0005) and IgM (r = 0.63, P < 0.0005) titers. Only 80% of IgG and 82% of IgM IFA readings determined by two technicians were within one dilution, while the DS was more reliable, with 100% within one dot. The rapidity, reliability, and simplicity of the DS suggest that it is a suitable test for use in clinical laboratories unable to perform the IFA test.

Measurement of CD4 T-lymphocyte levels is clinically useful in monitoring immune status in a number of conditions, including human immunodeficiency virus (HIV) infection, in which the absolute CD4 count is used to guide therapy. The absolute CD4 count is obtained by multiplying the results of the leukocyte count and the differential with a hematology cell counter and the percentage of CD4+ T lymphocytes determined by flow cytometry. These techniques require expensive, complex instrumentation, and interlaboratory results are difficult to standardize and reproduce. The rapid growth of HIV infection worldwide has increased the need for more-reproducible and cost-effective methods for CD4 T-cell monitoring. The TRAx CD4 test kit is based on a novel adaptation of conventional enzyme-linked immunosorbent assay (ELISA) and permits the simple quantitation of total CD4 protein from whole-blood lysates. In this study, the relationship between total CD4 protein measured in units per milliliter (TRAx) and in cells per microliter (flow cytometry and hematology) was defined in a multisite clinical study using linear regression analysis. Data from 230 HIV-seronegative and 321 HIV-seropositive specimens were used to calibrate the TRAx assay recombinant CD4 standards and controls in equivalent CD4 T lymphocytes per microliter (cells per microliter). The calibration of the TRAx CD4 assay in cells per microliter was validated with a second group of specimens from 17 healthy volunteers and 20 HIV-seropositive patients which were collected and tested under strictly controlled conditions intended to minimize the effects of specimen aging on the results of the reference method. These data were also used to estimate the variability of absolute CD4 count by cytometric methods as well as the precision of the TRAx CD4 result after it was calibrated in cells per microliter. Overall, correlations between the two methods ranged from 0.87 to 0.95. Additional studies demonstrated that the contribution of CD4 protein from monocytes and any soluble CD4 in sera are negligible in the TRAx assay and do not significantly affect results. This new method represents a promising alternative to absolute CD4 T-cell enumeration by flow cytometry and hematology.

The results of the DNA Analysis Module of the College of American Pathologists' Flow Cytometry Survey were analyzed to determine if participating laboratories could correctly identify aneuploid populations and estimate S-phase fraction reproducibly in proficiency specimens. Each survey contained three specimens: a diploid calibrator and two unknowns comprising cultured aneuploid tumor cells with DNA indexes of 1.2 to 2.0 admixed with normal lymphocytes. Results of ploidy analysis were obtained from a mean of 241 participants on 19 unknowns from 1990 to 1992. Most laboratories correctly reported the modal (and intended) number of aneuploid peaks; the proportion of correct responses ranged from 58% to 97%, with a mean of 86%. The lower percents of correct response were associated with near-diploid and tetraploid unknowns. Some laboratories showed consistent deviations from the mean for DNA index, indicating apparent instrument linearity problems. S-phase results on five specimens showed wide interlaboratory variability, although the means of participants' responses were close to the reference values obtained by the specimen provider.

Maintenance of CD4+ T helper lymphocyte counts has been used as a surrogate marker of efficacy for drugs in the treatment of AIDS. In a multicenter clinical trial, subtle improvement of CD4+ T cell counts may be masked and misinterpreted if care is not paid to likely sources that can contribute to the variability of measurement of CD4+ T lymphocytes. This review addresses major areas that can contribute to the variability of measurement of CD4+ T lymphocytes, with emphasis on applications to multicenter clinical trials, and proposes areas of improvement that may not be well recognized by the medical community. Whereas there are excellent guidelines for immunophenotyping, equal attention is needed in hematologic enumeration of WBC and absolute lymphocytes. In particular, allowing the margin of error acceptable to blood cell standards for HIV-infected specimens is unsatisfactory. Special attention should also be given to the stability of lymphocytes in the anticoagulant during storage, the lysing method, the quality assurance programs as well as intrasubject fluctuations which may be derived from exercise, medications and diurnal variations. Awareness of these contributing factors by physicians and technical analysts will expedite the discovery of potential therapy in the treatment of AIDS. For a multicenter clinical trial, it is advisable to select a centralized laboratory adopting a uniform protocol with regard to sample preparation and handling, using more stringent quality controls for hematologic analysers, calibration of instruments and immunophenotyping. Pending a true reference standard that can monitor the variation of the entire analytical procedure, we anticipate that future interlaboratory quality assurance programs will include absolute T lymphocyte count, an important parameter for assessing the accuracy and consistency of CD4+ T helper cell counts generated from a laboratory.

Objective: CD4 counts have been used to monitor progression of disease in HIV infection as criteria for initiation of therapy, and to stratify and follow patients in clinical trials. Recently, the Centers for Disease Control and Prevention (CDC) has made CD4 counts part of the classification of HIV disease. Because a CD4 percentage may be the only laboratory information available, this study was initiated to determine whether the correlation between CD4 percentages and CD4 counts is sufficiently high to enable these measures to be substituted for each other., Design, Setting and Patients: One thousand consecutive CD4 measurements from the University of Washington (UW) were used to create a model that was tested using datasets of 1000 CD4 measurements each from Maryland Medical Laboratories (MML) and Rush-Presbyterian-St Luke's Medical Center (Rush). The patients were not selected for age, sex, risk group or treatment. All patients from MML and Rush were known to be HIV-positive, while the HIV status of all UW patients was unknown., Results: The model predicted that a patient with a CD4 percentage > or = 14% would have a CD4 count > or = 200 x 10(6)/l(if CD4 percentage of 14% was used, 9% of patients would have a CD4 count > or = 200 x 10(6)/l), and a patient with a CD4 percentage > or = 27% would have a CD4 count > or = 500 x 10(6)/l(if CD4 percentage of 27% was used, 17% of patients would have a CD4 count > or = 500 x 10(6)/l)., Conclusions: These CD4 percentage correlations may be useful when a white blood cell and lymphocyte count are not available to calculate the CD4 count.