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While T cell lymphomas are classified as mature neoplasms, emerging evidence indicates that malignant transformation may occur at an earlier stage of T cell maturation. In this study, we determined clonal architectures in a broad range of T cell lymphomas. Our multidimensional profiling indicates that a large part of these lymphomas in fact emerge from an immature lymphoid T cell precursor at a maturation stage prior to V(D)J rearrangement that undergoes branching evolution. Consequently, at single cell resolution we observed considerable clonal tiding under selective therapeutic pressure. T cell receptor next-generation sequencing suggested a highly biased usage of TRBV20-1 gene segments as part of multiple antigen receptor rearrangements per patient. The predominance of TRBV20-1 was found across all major T cell lymphoma subtypes analyzed. This suggested that this particular V gene – independently of complementarity-determining region 3 (CDR3) configuration - may represent a driver of malignant transformation. Together, our data indicate that T cell lymphomas derive from immature lymphoid precursors and display considerable intratumoral heterogeneity that may provide the basis for relapse and resistance in these hard-to-treat cancers.

The classification of B cell lymphomas-mainly based on light microscopy evaluation by a pathologist-requires many years of training. Since the B cell receptor (BCR) of the lymphoma clonotype and the microenvironmental immune architecture are important features discriminating different lymphoma subsets, we asked whether BCR repertoire next-generation sequencing (NGS) of lymphoma-infiltrated tissues in conjunction with machine learning algorithms could have diagnostic utility in the subclassification of these cancers. We trained a random forest and a linear classifier via logistic regression based on patterns of clonal distribution, VDJ gene usage and physico-chemical properties of the top-n most frequently represented clonotypes in the BCR repertoires of 620 paradigmatic lymphoma samples-nodular lymphocyte predominant B cell lymphoma (NLPBL), diffuse large B cell lymphoma (DLBCL) and chronic lymphocytic leukemia (CLL)-alongside with 291 control samples. With regard to DLBCL and CLL, the models demonstrated optimal performance when utilizing only the most prevalent clonotype for classification, while in NLPBL-that has a dominant background of non-malignant bystander cells-a broader array of clonotypes enhanced model accuracy. Surprisingly, the straightforward logistic regression model performed best in this seemingly complex classification problem, suggesting linear separability in our chosen dimensions. It achieved a weighted F1-score of 0.84 on a test cohort including 125 samples from all three lymphoma entities and 58 samples from healthy individuals. Together, we provide proof-of-concept that at least the 3 studied lymphoma entities can be differentiated from each other using BCR repertoire NGS on lymphoma-infiltrated tissues by a trained machine learning model.

Background: Uroflowmetry (UF) is an established procedure in urology and is recommended before further investigations of neurogenic lower urinary tract dysfunction (NLUTD). Some authors even consider using UF instead of urodynamics (UD). Studies on the interrater reliability of UF regarding treatment recommendations are rare, and there are no relevant data on people with multiple sclerosis (PwMS). The aim of this study was to investigate the interrater reliability (IRR) of UF concerning diagnosis and therapy in PwMS prospectively. Methods: UF of 92 PwMS were assessed by 4 raters. The diagnostic criteria were normal findings (NFs), detrusor overactivity (DO), detrusor underactivity (DU), detrusor–sphincter dyssynergia (DSD) and bladder outlet obstruction (BOO). The possible treatment criteria were as follows: no treatment (NO), catheter placement (CAT), alpha-blockers, detrusor-attenuating medication, botulinum toxin (BTX), neuromodulation (NM), and physiotherapy/biofeedback (P/BF). IRR was assessed by kappa (κ). Results: κ of diagnoses were NFs = 0.22; DO = 0.17; DU = 0.07; DSD = 0.14; and BOO = 0.18. For therapies, the highest κ was BTX = 0.71, NO = 0.38 and CAT = 0.44. Conclusions: There is a high influence of the individual rater. UD should be subject to the same analysis and a comparison should be made between UD and UF. This may have implications for the value of UF in the neuro-urological management of PwMS, although at present UD remains the gold standard for the diagnostics of NLUTD in PwMS.

Background Broad-scale monitoring of arthropods is often carried out with passive traps (e.g., Malaise traps) that can collect thousands of specimens per sample. The identification of individual specimens requires time and taxonomic expertise, limiting the geographical and temporal scale of research and monitoring studies. DNA metabarcoding of bulk-sample homogenates has been found to be faster, efficient and reliable, but the destruction of samples prevents a posteriori validation of species occurrences and relative abundances. Non-destructive metabarcoding of DNA extracted from collection medium has been applied in a limited number of studies, but further tests of efficiency are required with different trap types and collection media to assess the consistency of the method. Methods We quantified the detection rate of arthropod species when applying non-destructive DNA metabarcoding with a short (127-bp) fragment of mitochondrial COI on two combinations of passive traps and collection media: (1) water with monopropylene glycol (H2O–MPG) used in window-flight traps (WFT, 53 in total); (2) ethanol with monopropylene glycol (EtOH–MPG) used in Malaise traps (MT, 27 in total). We then compared our results with those obtained for the same samples using morphological identification (for WFTs) or destructive metabarcoding of bulk homogenate (for MTs). This comparison was applied as part of a larger study of arthropod species richness in silver fir (Abies alba Mill., 1759) stands across a range of climate-induced tree dieback levels and forest management strategies. Results Of the 53 H2O-MPG samples from WFTs, 16 produced no metabarcoding results, while the remaining 37 samples yielded 77 arthropod MOTUs in total, of which none matched any of the 343 beetle species morphologically identified from the same traps. Metabarcoding of 26 EtOH–MPG samples from MTs detected more arthropod MOTUs (233) than destructive metabarcoding of homogenate (146 MOTUs, 8 orders), of which 71 were shared MOTUs, though MOTU richness per trap was similar between treatments. While we acknowledge the failure of metabarcoding from WFT-derived collection medium (H2O–MPG), the treatment of EtOH-based Malaise trapping medium remains promising. We conclude however that DNA metabarcoding from collection medium still requires further methodological developments and cannot replace homogenate metabarcoding as an approach for arthropod monitoring. It can be used nonetheless as a complementary treatment when enhancing the detection of soft-bodied arthropods like spiders and Diptera.

Fluctuating environments threaten fertility and viability. To better match the immediate, local environment, many organisms adopt alternative phenotypic states, a phenomenon called "phenotypic plasticity." Natural populations that predictably encounter fluctuating environments tend to be more plastic than conspecific populations that encounter a constant environment, suggesting that phenotypic plasticity can be adaptive. Despite pervasive evidence of such "adaptive phenotypic plasticity," gene regulatory mechanisms underlying plasticity remains poorly understood. Here we test the hypothesis that environment-dependent phenotypic plasticity is mediated by epigenetic factors. To test this hypothesis, we exploit the adaptive reproductive arrest of Drosophila melanogaster females, called diapause. Using an inbred line from a natural population with high diapause plasticity, we demonstrate that diapause is determined epigenetically: only a subset of genetically identical individuals enter diapause and this diapause plasticity is epigenetically transmitted for at least three generations. Upon screening a suite of epigenetic marks, we discovered that the active histone marks H3K4me3 and H3K36me1 are depleted in diapausing ovaries. Using ovary-specific knockdown of histone mark writers and erasers, we demonstrate that H3K4me3 and H3K36me1 depletion promotes diapause. Given that diapause is highly polygenic, that is, distinct suites of alleles mediate diapause plasticity across distinct genotypes, we also investigated the potential for genetic variation in diapause-determining epigenetic marks. Specifically, we asked if these histone marks were similarly depleted in diapause of a genotypically distinct line. We found evidence of divergence in both the gene expression program and histone mark abundance. This study reveals chromatin determinants of phenotypic plasticity and suggests that these determinants may be genotype-dependent, offering new insight into how organisms may exploit and evolve epigenetic mechanisms to persist in fluctuating environments.

A female in her 60s presented to the allergy and immunology clinic for further investigation of ongoing dermatitis. She presented with chronic acrocyanosis, mainly in her left lower extremity, extending distally from her mid thigh with concurrent ulcerations in her foot resulting in immobility secondary to pain. She experienced these symptoms for years without a definitive diagnosis. The lack of diagnosis was due, in part, to her atypical symptoms and laboratory findings that required a high level of clinical suspicion to diagnose. Extensive autoimmune workup was largely unrevealing with the exception of a cold agglutinin titer of 1:250 and a positive anticomplement C3b direct antiglobulin test. A diagnosis of cold agglutinin disease was made and treatment with rituximab monotherapy was initiated.

ABSTRACT: Mechanical loading has been used increasingly in recent years alongside the manual loading of broilers. The aim of this study was to analyze the influence of various factors on the behavior of broilers and the impacts occurring to broilers during loading with a loading machine in order to identify risk factors and thus improve animal welfare. During 32 loadings we assessed escape behavior, wing flapping, flip, bumping into an animal, and bumping against the machine or container by evaluation of video recordings. The parameters were analyzed for influences of rotation speed, container type (GP container vs. SmartStack container), husbandry system (“Indoor Plus” vs. “Outdoor Climate”), and season. In addition, the behavior and impact parameters were correlated with the loading-related injuries. The results showed that an increase in rotation and conveyor belt speed was associated with an elevated risk of all behaviors or impacts, except for a lower risk of escape behavior. The seasonal influence showed that the risk of wing flapping, bumping into an animal, and bumping against the machine or container was highest in fall. The comparison of the container types showed that the risk of escape behavior, wing flapping, and bumping into an animal was increased when loading with the SmartStack container, but the risk of bumping against the machine or container was reduced. For animals of the husbandry system “Outdoor Climate,” the risk of bumping into an animal and bumping against the machine or container was reduced. In addition, we found an influence of the examined parameters on the loading-related injuries. Increased escape behavior reduced the risk of severe injuries (fractures, dislocations, epiphysiolyses). Wing flapping and bumping against the machine or container increased the risk of hematomas and abrasions. The risk of hematomas was increased if broilers bumped into conspecifics. In summary, our analysis revealed that the behavior and impacts on the animals during loading are influenced by each of the analyzed factors and that they in turn can affect loading-related injuries.

Sire et al. examine the effects of climate-induced forest dieback and salvage logging on insect diversity in silver fir Pyrenean forests using metabarcoding. Although no consistent variation in species richness was found, forest dieback cause massive changes in community composition particularly affecting rare species.

Background: Lesions in the periventricular, (juxta)cortical, and infratentorial region, as visible on brain MRI, are part of the diagnostic criteria for Multiple sclerosis (MS) whereas lesions in the subcortical region are currently only a marker of disease activity. It is unknown whether MS lesions follow individual spatial patterns or whether they occur in a random manner across diagnostic regions. Aim: First, to describe cross-sectionally the spatial lesion patterns in patients with MS. Second, to investigate the spatial association of new lesions and lesions at baseline across diagnostic regions. Methods: Experienced neuroradiologists analyzed brain MRI (3D, 3T) in a cohort of 330 early MS patients. Lesions at baseline and new solitary lesions after two years were segmented (manually and by consensus) and classified as periventricular, (juxta)cortical, or infratentorial (diagnostic regions) or subcortical—with or without Gadolinium-enhancement. Gadolinium enhancement of lesions in the different regions was compared by Chi square test. New lesions in the four regions served as dependent variable in four zero-inflated Poisson models each with the six independent variables of lesions in the four regions at baseline, age and gender. Results: At baseline, lesions were most often observed in the subcortical region (mean 13.0 lesions/patient), while lesion volume was highest in the periventricular region (mean 2287 µl/patient). Subcortical lesions were less likely to show gadolinium enhancement (3.1 %) than juxtacortical (4.3 %), periventricular (5.3 %) or infratentorial lesions (7.2 %). Age was inversely correlated with new periventricular, juxtacortical and subcortical lesions. New lesions in the periventricular, juxtacortical and infratentorial region showed a significant autocorrelative behavior being positively related to the number of lesions in the respective regions at baseline. New lesions in the subcortical region showed a different behavior with a positive association with baseline periventricular lesions and a negative association with baseline infratentorial lesions. Conclusion: Across regions, new lesions do not occur randomly; instead, new lesions in the periventricular, juxtacortical and infratentorial diagnostic region are associated with that at baseline. Lesions in the subcortical regions are more closely related to periventricular lesions. Moreover, subcortical lesions substantially contribute to lesion burden in MS but are less likely to show gadolinium enhancement (than lesions in the diagnostic regions).

ABSTRACT: The aim of this study was to compare the 2 manual catching methods in terms of injuries and behavior. Throughout 12 loadings on practical farms with the same standard, 1 container each was caught using the one-legged (1LCM) and 1 using the two-legged catching method (2LCM). The animals were filmed during loading to evaluate their behavior and subsequently examined regarding injuries. Wing flapping was observed more frequently in broilers caught with the 1LCM than 2 LCM. Carrying animals with neighbors (1 neighbor: P < 0.001; 2 neighbors: P < 0.001) and a grasping position at or above the tarsal joint (P < 0.001; P < 0.054) reduced wing flapping in both methods. A short grasping duration (P = 0.004), settling the broilers into the crate (P = 0.005) and avoiding striking the broilers against the crate (P < 0.001) reduced the occurrence of wing flapping. About 1.1% of 1LCM and 0.43% of 2LCM broilers were diagnosed with an epiphysiolysis. Catching with the 1LCM (P = 0.042), loading in lower crates (low vs. middle: P = 0.005; low vs. high: P = 0.008), a longer catching duration (p = 0.025) and female broilers (P = 0.007) had a higher chance for epiphysiolysis. Broilers loaded in lower crates (P = 0.007) and ones which showed more wing flapping (P = 0.015) had a higher chance for hematomas. A higher loading duration led to a higher risk of hematomas (prevalence: 1.5%) and a prevalence of 1.0% of broilers with severe injury in 2LCM in a simultaneously performed study (mechanical loading vs. 2LCM), in which manual loadings of entire barns were evaluated. This may be caused by fatigue of the workers. In summary, the catching method or number of grasped legs is not the decisive factor, but the compliance and implementation of the identified risks and careful handling of the animals are determining factors to reduce wing injuries caused by loading and wing flapping.

ABSTRACT: Various plumage and integument scoring methods are commonly used to deduce the occurrence of severe feather pecking and cannibalism in laying hens. The aim of our study was to provide evidence of correlations between the occurrence of severe feather pecking and our individual plumage scoring system used under practical conditions on commercial farms with non–beak-trimmed and beak-trimmed layers (study I). In second step, we aimed to verify whether the results of the elaborate individual scoring may be predicted with a visual scoring method based on the total body scores of groups of birds (study II). For study I we observed the pecking behavior and performed an individual plumage scoring at the beginning, in the middle, and at the end of a laying period on 8 commercial farms. For study II we performed both an individual and a visual plumage scoring on 49 flocks on 45 farms at the beginning of the laying period and on 43 flocks on 41 farms at the end of the laying period. Spearman's Rho revealed a correlation of the mean feather pecking rate with the total plumage score, the neck–back plumage score, and the total cannibalism score in all observation periods. A high feather pecking rate was correlated with severe plumage damage and the frequent occurrence of skin injuries. We conclude that both the total plumage score and the neck–back plumage score constitute a reliable indicator of the occurrence of severe feather pecking in the flocks assessed in this study. The results of study II suggest that the percental assessment of plumage damage on flock level in 3 categories (“visual score”) leads to a good prognosis of the actual, individually assessed plumage score. Therefore, the application (and documentation) of the visual score on a regular basis can provide a good evaluation of the development of the plumage condition of the flock. The visual score presented in this study is suggested as a suitable instrument for self-evaluation programs on farms.

ABSTRACT: This study aimed to develop and validate a camera vision score that could detect macroscopic alterations of the hock, to identify errors and to assess possible factors that could influence the assessment. Two hundred hocks in the first (calibration) phase and 500 hocks in the second (validation) phase were collected at slaughter, visually assessed, placed back into the evisceration line and assessed by a camera system with 2 software systems. The size of the alteration in percent (%) measured by the camera system was evaluated (“camera score”, CS). Additionally, temperature, humidity, and light intensities were measured. In the calibration phase, threshold values of camera scores for respective macro scores were defined and performance measures evaluated. In the validation phase, the generated threshold values were validated, occurring errors, as well as possible impacts of climatic factors analyzed. The results showed that the generated thresholds predict the camera score values at which the respective macro score has the highest probability of appearance. Small hock burn lesions ≤0.5 cm have the highest probability at a camera score of ≥0.2 (original CS) or ≥0.1 (updated CS), and lesions >0.5 cm have the highest probability at a camera score of ≥0.7 (original CS) or ≥1.1 (updated CS). Large lesions (>0.5 cm) are more reliably identified by the system than small lesions. The risks of errors in assessing reference areas and lesions showed a correct identification of lesions to be the most probable result even if the reference area is not correctly identified. The probability of a correct identification of lesions by the camera system was slightly higher (not significant) with the updated software (risk = 0.66 [0.62–0.70]) than with the original software (risk = 0.63 [0.58–0.67]). Automatic assessment systems at slaughter could be adjusted to the presented threshold values to classify hock burn lesions. Software adaptations can improve the performance measures of diagnosis and reduce the probability of errors.

Loading of broilers for transport to the processing plant poses a notable injury risk for broilers. Therefore, the poultry industry has developed mechanical methods as alternatives to manual loading methods. Our objective in the present study was to compare manual loading (MAN) of broilers with the mechanical loading (MECH). We assessed the injuries of broilers of 12 MAN and 12 MECH flocks on-farm before and immediately after loading, documented the numbers of broilers dead on arrival reported by the processing plant, and assessed the circumstances at loading. A smaller number of broilers with a hematoma (≥0.5 cm in diameter) on the wing were observed after MAN compared with MECH using the examined harvester (MAN vs. MECH odds ratio: 0.16; 95% confidence interval: 0.10, 0.28). The number of broilers with severe wing injuries did not differ between the loading methods. The number of broilers dead on arrival was greater in mechanically loaded flocks (MAN vs. MECH odds ratio: 0.26; 95% confidence interval: 0.10, 0.68), but lower than in comparable studies. We observed a lower average stocking rate than targeted in the drawers of MECH containers, most likely because the used harvester can adapt to short-term changes in weight and adjust the stocking rate during the loading process. A longer total loading duration in MAN was associated with an increase of wing hematomas, and the involvement of more working people per 10,000 broilers during MAN was associated with a lower occurrence of hematomas. The total loading duration in MECH had no notable influence on the occurrence of injuries. Physical conditions of the involved personnel might play a larger role in MAN than in MECH. The harvester that was examined should be further developed to reduce the occurrence of hematomas. Our results indicate that the choice of loading method alone does not determine the injury risk, and multiple factors are associated with broiler welfare during loading. It is important that the chosen method is performed under the most adequate conditions.

Abstract Background The Cumberland Gap Region (CGR) of the United States is a natural corridor between the southeastern, northeastern, and midwestern regions of the country. CGR has also many species of ticks and mosquitos that serve as competent vectors for important animal and human pathogens. In this study, we tested dogs from six different animal shelters in the CGR for Rocky Mountain spotted fever (RMSF), anaplasmosis, Lyme disease, canine ehrlichiosis and canine heartworm disease. Results Sera from 157 shelter dogs were tested for antibodies to RMSF agent, Rickettsia rickettsii, using an indirect immunofluorescence assay. Sixty-six dogs (42.0%) were positive for either IgM or IgG, or both IgM and IgG antibodies to R. rickettsii. Moreover, the same set of sera (n = 157) plus an and additional sera (n = 75) from resident dogs at the same shelters were tested using the SNAP 4Dx Plus. Of 232 dogs tested, two (0.9%) were positive for antibodies to Anaplasma phagocytophilum/A. platys, nine (3.9%) were positive for antibodies to Borrelia burgdorferi, 23 (9.9%) for positive for antibodies to Ehrlichia canis/E. ewingii, and 13 (5.6%) were positive for Dirofilaria immitis antigen. Co-infection with two or more etiologic agents was detected in five animals. Three dogs had antibodies to both B. burgdorferi and E. canis/E. ewingii, and two dogs were positive for D. immitis antigen and antibodies to B. burgdorferi and E. canis/E. ewingii. Conclusions Shelter dogs in the CGR are exposed to a number of important vector-borne pathogens. Further studies are required to ascertain the roles these animals play in maintenance and transmission of these pathogens.

Introduction Sepsis is a common, potentially life-threatening complication of infection. The optimal treatment for sepsis includes prompt antibiotics and intravenous fluids, facilitated by its early and accurate recognition. Currently, clinicians identify and assess severity of suspected sepsis using validated clinical scoring systems. In England, the National Early Warning Score 2 (NEWS2) has been mandated across all National Health Service (NHS) trusts and ambulance organisations. Like many clinical scoring systems, NEWS2 should not be used without clinical judgement to determine either the level of acuity or a diagnosis. Despite this, there is a tendency to overemphasise the score in isolation in patients with suspected infection, leading to the overprescription of antibiotics and potentially treatment-related complications and rising antimicrobial resistance. The biomarker procalcitonin (PCT) has been shown to be useful in specific circumstances to support appropriate antibiotics prescribing by identifying bacterial infection. PCT is not routinely used in the care of undifferentiated patients presenting to emergency departments (EDs), and the evidence base of its optimal usage is poor. The PROcalcitonin and NEWS2 evaluation for Timely identification of sepsis and Optimal (PRONTO) study is a randomised controlled trial (RCT) in adults with suspected sepsis presenting to the ED to compare standard clinical management based on NEWS2 scoring plus PCT-guided risk assessment with standard clinical management based on NEWS2 scoring alone and compare if this approach reduces prescriptions of antibiotics without increasing mortality.Methods and analysis PRONTO is a parallel two-arm open-label individually RCT set in up to 20 NHS EDs in the UK with a target sample size of 7676 participants. Participants will be randomised in a ratio of 1:1 to standard clinical management based on NEWS2 scoring or standard clinical management based on NEWS2 scoring plus PCT-guided risk assessment. We will compare whether the addition of PCT measurement to NEWS2 scoring can lead to a reduction in intravenous antibiotic initiation in ED patients managed as suspected sepsis, with at least no increase in 28-day mortality compared with NEWS2 scoring alone (in conjunction with local standard care pathways). PRONTO has two coprimary endpoints: initiation of intravenous antibiotics at 3 hours (superiority comparison) and 28-day mortality (non-inferiority comparison). The study has an internal pilot phase and group-sequential stopping rules for effectiveness and futility/safety, as well as a qualitative substudy and a health economic evaluation.Ethics and dissemination The trial protocol was approved by the Health Research Authority (HRA) and NHS Research Ethics Committee (Wales REC 2, reference 20/WA/0058). In England and Wales, the law allows the use of deferred consent in approved research situations (including ED studies) where the time dependent nature of intervention would not allow true informed consent to be obtained. PRONTO has approval for a deferred consent process to be used. Findings will be disseminated through peer-reviewed journals and presented at scientific conferences.Trial registration number ISRCTN54006056.

Surgical castration of piglets is generally recognized as a painful procedure, but there is currently no gold standard for the assessment of pain behavior in piglets. However, pain assessment is essential for evaluating the effectiveness of local anesthetics. In this study, we investigated the efficacy of four local anesthetics in terms of pain relief during and after surgical castration in three sequential study parts. To do so, we filmed 178 piglets before the applied procedures, after injection of the local anesthetic, and up to 24 h after castration (five observation times in total) in an observation arena and compared their behavior before and after castration and between treatments and control groups. The results showed significant differences in the behavior of the piglets before and after castration and between the sham-castrated control group and the control group castrated without anesthesia. The different local anesthesia treatment groups showed diverging differences to the control groups. The most frequently shown pain-associated behaviors of the piglets were changes in tail position and hunched back posture. We observed a reduction but no complete elimination of the expressed pain-associated behaviors after local anesthesia. Several behavioral changes—such as changes in tail position, hunched back posture or tail wagging—persisted until the day after castration. Owing to the limited duration of the effects of the local anesthetics, local anesthesia did not influence long-term pain.

Evolve-and-resequence (E+R) experiments leverage next-generation sequencing technology to track the allele frequency dynamics of populations as they evolve. While previous work has shown that adaptive alleles can be detected by comparing frequency trajectories from many replicate populations, this power comes at the expense of high-coverage (>100x) sequencing of many pooled samples, which can be cost-prohibitive. Here, we show that accurate estimates of allele frequencies can be achieved with very shallow sequencing depths (

To advance our understanding of adaptation to temporally varying selection pressures, we identified signatures of seasonal adaptation occurring in parallel among Drosophila melanogaster populations. Specifically, we estimated allele frequencies genome-wide from flies sampled early and late in the growing season from 20 widely dispersed populations. We identified parallel seasonal allele frequency shifts across North America and Europe, demonstrating that seasonal adaptation is a general phenomenon of temperate fly populations. Seasonally fluctuating polymorphisms are enriched in large chromosomal inversions, and we find a broad concordance between seasonal and spatial allele frequency change. The direction of allele frequency change at seasonally variable polymorphisms can be predicted by weather conditions in the weeks prior to sampling, linking the environment and the genomic response to selection. Our results suggest that fluctuating selection is an important evolutionary force affecting patterns of genetic variation in Drosophila.

The objective of this study was to describe the ontogeny of the severe feather pecking (SFP), gentle feather pecking (GFP), aggressive pecking (AP), and enrichment pecking (EP) of non-beak-trimmed Lohmann Brown (LB)-pullets during the first 4 weeks of life (observation on 1st, 8th, 15th, 22nd, and 29th days of life) while they were kept within the compartments of a commercial rearing aviary (without access to a litter area). All chicks were placed into compartments of the middle tier of the aviary on the 1st day of life. On the 10th day of life, half of the chicks of each compartment were moved into the compartments of the lower tier. The aviary floor was covered with chick paper before the placement of the chicks and fully or partially removed from the 15th day of life onwards. The pecking behavior patterns were observed in three experimental groups (EG): NE (not enriched): group size until/after 10th day of life: 230/115; 120.8/60.4 birds/m2, no enrichment; EL (enriched, low stocking rate): group size until/after 10th day of life: 203/101, 106.6/53.6 birds/m2; and EH (enriched, high stocking rate): group size until/after 10th day of life: 230/115;120.8/60.4 birds/m2, both pecking stones and blocks as enrichment) in two rearing periods. For each pecking behavior pattern, an independent regression model with the parameters EG, chick paper, observation day, and functional area was estimated. GFP, SFP, and EP increased with age during the observation. The AP rate was highest in all EGs on the first day of life and decreased during the observation period. A pairwise comparison of NE (high stocking rate without enrichment) with EH (high stocking rate with enrichment and with EL (low stocking rate with enrichment) showed a significant effect of the EG on pecking behavior, with more SFP, AP, and GFP in NE. There were no differences between EL and EH, indicating that the provision of pecking materials had more influence than the stocking rate. However, we presumed that the difference between the stocking rates were too small to observe an effect. AP, SFP, and GFP were significantly higher on wired slats, as compared to the perches and the vicinity of the enrichment materials. The enrichment materials were suitable and intensely used by the pullets. The provision of pecking blocks and pecking stones was recommended as a preventive measure from the first day of life onwards for pullets housed in commercial rearing aviaries. There was no effect of reduced stocking rate, most likely due to the low variation in stocking rates.

Organisms living in seasonally variable environments utilize cues such as light and temperature to induce plastic responses, enabling them to exploit favorable seasons and avoid unfavorable ones. Local adapation can result in variation in seasonal responses, but the genetic basis and evolutionary history of this variation remains elusive. Many insects, including Drosophila melanogaster, are able to undergo an arrest of reproductive development (diapause) in response to unfavorable conditions. In D. melanogaster, the ability to diapause is more common in high latitude populations, where flies endure harsher winters, and in the spring, reflecting differential survivorship of overwintering populations. Using a novel hybrid swarm-based genome wide association study, we examined the genetic basis and evolutionary history of ovarian diapause. We exposed outbred females to different temperatures and day lengths, characterized ovarian development for over 2800 flies, and reconstructed their complete, phased genomes. We found that diapause, scored at two different developmental cutoffs, has modest heritability, and we identified hundreds of SNPs associated with each of the two phenotypes. Alleles associated with one of the diapause phenotypes tend to be more common at higher latitudes, but these alleles do not show predictable seasonal variation. The collective signal of many small-effect, clinally varying SNPs can plausibly explain latitudinal variation in diapause seen in North America. Alleles associated with diapause are segregating in Zambia, suggesting that variation in diapause relies on ancestral polymorphisms, and both pro- and anti-diapause alleles have experienced selection in North America. Finally, we utilized outdoor mesocosms to track diapause under natural conditions. We found that hybrid swarms reared outdoors evolved increased propensity for diapause in late fall, whereas indoor control populations experienced no such change. Our results indicate that diapause is a complex, quantitative trait with different evolutionary patterns across time and space.

The assessment of foot pad dermatitis at slaughter is a suitable method to assess and monitor the welfare of broilers. The goals of this study were to define and validate a camera-based score that could identify macroscopic lesions of the foot pads, to identify errors, and to assess possible external factors that could influence the assessment. In the first phase 200 feet of broilers and in the second phase 500 feet were collected at slaughter, assessed visually, hung back into the evisceration line, and assessed by an automatic system. The camera score cut-off values were defined in the first (=calibration) phase. In the second (=validation) phase, the performance of diagnosis for these cut-off values was evaluated, and possible errors in the assessment of reference surface area and foot pad lesions were analyzed. The results showed that, in particular, Macro Scores 0, 2, and 3 could be identified with sufficiently high sensitivity. For Macro Score 1, the sensitivity of diagnosis was not sufficiently high in the two evaluated software versions. The current automatic assessment systems at slaughter could be adjusted to the cut-off values in order to classify foot pad dermatitis lesions. Furthermore, software updates can enhance the performance measures and lower the probability of errors.

Background: Multiple sclerosis patients often develop neurogenic lower urinary tract dysfunction with a potential risk of upper urinary tract damage. Diagnostic tools are urodynamics, bladder diary, uroflowmetry, and post-void residual, but recommendations for their use are controversial. Objective: We aimed to identify clinical parameters indicative of neurogenic lower urinary tract dysfunction in multiple sclerosis patients. Methods: 207 patients were prospectively assessed independent of the presence of lower urinary tract symptoms. We analyzed Expanded Disability Status Scale scores, uroflowmetry, post-void residual, rate of urinary tract infections, standardized voiding frequency, and voided volume in correlation with urodynamic findings. Results: We found a significant correlation between post-void residual (odds ratio (OR) 4.17, confidence interval (CI) 1.20–22.46), urinary tract infection rate (OR 3.91, CI 1.13–21.0), voided volume (OR 4.53, CI 1.85–11.99), increased standardized voiding frequency (OR 7.40, CI 2.15–39.66), and urodynamic findings indicative of neurogenic lower urinary tract dysfunction. Expanded Disability Status Scale shows no correlation. Those parameters (except post-void residual) are also associated with reduced bladder compliance, as potential risk for kidney damage. Conclusion: Therefore, bladder diary and urinary tract infection rate should be routinely assessed to identify patients who require urodynamics.

Abstract Background Universities are increasingly implementing programs to effectively respond to and manage sport-related concussions (SRCs). One such effort is to develop common data elements (CDEs) and standardize data collection methods. The objectives of this study were to describe CDEs currently collected by Big Ten and Ivy League universities for SRC studies, and to compare the data collected with the core CDEs recommended by the National Institute of Neurological Disorders and Stroke (NINDS). Methods We conducted an anonymous cross-sectional online survey among medical staff at the 14 Big Ten and 8 Ivy League universities (one per university) between September and October 2015. The survey instrument, including 9 questions corresponding to the concussion data collected before, during, and after a concussion, was developed and pilot-tested before field use. We analyzed patterns of the concussion CDEs being collected, including when, what, and how the data were collected and stored, and compared them with the NINDS' recommended core CDEs. Results A total of 19 out of 22 universities were included, with 13 from Big Ten and 6 from Ivy-League universities. All 19 participating universities currently collected concussion data with athletes before, during, and after a concussion. Great similarities in data collection were observed at baseline and acutely post-concussion across participating universities. All 19 universities collected at least one of the ten recommended acute symptoms checklists, and 18 universities collected one of the four recommended core neuropsychological function cognitive measures. However, CDEs in the sub-acute and chronic timeframes were limited, with only 9 (47%) universities collecting post-concussion short to long term outcome data. While over 60% of universities collected and stored concussion data electronically, only 17% to 42% of data collected were readily available for research. Conclusions Significant inter-institutional similarities in acute concussion CDEs were found. Further efforts should focus on collecting sub-acute and chronic timeframe core CDEs and creating data access protocols to facilitate evidence-based concussion prevention and treatment for all collegiate athletes.

Abstract Background and purpose New or enlarging T2‐hyperintense white matter lesions (WML) are associated with clinical disease progression in multiple sclerosis (MS). The prognostic value of WML shrinking is unclear. Assuming that waning of acute inflammation and repair processes would be the main drivers of WML shrinking, we aimed to assess the prognostic value of WML shrinking in early MS. Methods We retrospectively analyzed a cohort of 144 early MS patients with three brain MRI scans at baseline and after 1 and 3 years available. All patients were therapy naïve at baseline and 70.5% of them treated with disease modifying drugs at year 1. We determined the volume of WML shrinking between MRI scans, total WML volumes, number of gadolinium‐enhancing and new WML, white matter (WM) and gray matter volumes at each MRI scan. Clinical disability was measured by Expanded Disability Status Scale. We performed the correlation analyses of WML shrinking with other MRI parameters and clinical outcome. Results White matter lesions shrinking was highly variable between patients and correlated with the initial number of gadolinium‐enhancing WML and with WM volume decrease. WML shrinking was not associated with clinical outcome. Conclusion We found no indication of a prognostic value of WML shrinking in early MS patients. WML shrinking seems to be related to waning of acute inflammation.

Objectives Omissions and delays in delivering nursing care are widely reported consequences of staffing shortages, with potentially serious impacts on patients. However, studies so far have relied almost exclusively on nurse self-reporting. Monitoring vital signs is a key part of nursing work and electronic recording provides an opportunity to objectively measure delays in care. This study aimed to determine the association between registered nurse (RN) and nursing assistant (NA) staffing levels and adherence to a vital signs monitoring protocol.Design Retrospective observational study.Setting 32 medical and surgical wards in an acute general hospital in England.Participants 538 238 nursing shifts taken over 30 982 ward days.Primary and secondary outcome measures Vital signs observations were scheduled according to a protocol based on the National Early Warning Score (NEWS). The primary outcome was the daily rate of missed vital signs (overdue by ≥67% of the expected time to next observation). The secondary outcome was the daily rate of late vital signs observations (overdue by ≥33%). We undertook subgroup analysis by stratifying observations into low, medium and high acuity using NEWS.Results Late and missed observations were frequent, particularly in high acuity patients (median=44%). Higher levels of RN staffing, measured in hours per patient per day (HPPD), were associated with a lower rate of missed observations in all (IRR 0.983, 95% CI 0.979 to 0.987) and high acuity patients (0.982, 95% CI 0.972 to 0.992). However, levels of NA staffing were only associated with the daily rate (0.954, CI 0.949 to 0.958) of all missed observations.Conclusions Adherence to vital signs monitoring protocols is sensitive to levels of nurse and NA staffing, although high acuity observations appeared unaffected by levels of NAs. We demonstrate that objectively measured omissions in care are related to nurse staffing levels, although the absolute effects are small.Study registration The data and analyses presented here were part of the larger Missed Care study (ISRCTN registration: 17930973).

Feather pecking and cannibalism are behavioral disorders that cause animal-welfare-relevant and economic problems. To mitigate these problems, the beaks of conventionally reared turkeys are usually already trimmed in the hatcheries. To find an alternative to beak trimming, we conducted this study with male turkeys of three breeds: B.U.T. 6, B.U.T. Premium and, Auburn (200 turkeys per breed). Half of the birds had infrared-trimmed beaks; the other half had intact beaks. For each treatment combination (breed, beak status), 25 turkeys were housed in one section. A screed grinding wheel was installed in each feed pan of the non-beak-trimmed turkeys as of week six to facilitate natural beak abrasion until slaughter. Eight randomly selected turkeys per section were regularly examined to record injuries, plumage condition, and beak dimensions. In addition, 96 beaks from randomly slaughtered birds were examined macroscopically and histologically. The results concerning injuries and plumage condition showed in most cases no differences between the beak-trimmed turkeys and the ones provided with the blunting disks. The histological examinations revealed alterations in only the beak-trimmed birds. We can conclude that the blunting method smoothens the beak during feeding and thus may be a possible alternative to beak trimming.

Immune-related adverse events have been reported in patients treated with anti-programmed death-1 receptor drugs such as nivolumab. We present a case of a new-onset seronegative rheumatoid arthritis in a patient with metastatic melanoma treated with nivolumab.

Longitudinal analysis of white matter lesion changes on serial MRI has become an important parameter to study diseases with white-matter lesions. Here, we build on earlier work on cross-sectional lesion segmentation; we present a fully automatic pipeline for serial analysis of FLAIR-hyperintense white matter lesions. Our algorithm requires three-dimensional gradient echo T1- and FLAIR- weighted images at 3 Tesla as well as available cross-sectional lesion segmentations of both time points. Preprocessing steps include lesion filling and intrasubject registration. For segmentation of lesion changes, initial lesion maps of different time points are fused; herein changes in intensity are analyzed at the voxel level. Significance of lesion change is estimated by comparison with the difference distribution of FLAIR intensities within normal appearing white matter. The method is validated on MRI data of two time points from 40 subjects with multiple sclerosis derived from two different scanners (20 subjects per scanner). Manual segmentation of lesion increases served as gold standard. Across all lesion increases, voxel-wise Dice coefficient (0.7) as well as lesion-wise detection rate (0.8) and false-discovery rate (0.2) indicate good overall performance. Analysis of scans from a repositioning experiment in a single patient with multiple sclerosis did not yield a single false positive lesion. We also introduce the lesion change plot as a descriptive tool for the lesion change of individual patients with regard to both number and volume. An open source implementation of the algorithm is available at http://www.statistical-modeling.de/lst.html. Keywords: Magnetic resonance imaging, Multiple sclerosis, White matter lesions, Lesion segmentation

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system with presumed autoimmune origin. The development of lesions within the gray matter and white matter, which are highly variable with respect to number, total volume, morphology and spatial evolution and which only show a limited correlation with clinical disability, is a hallmark of the disease. Population-based studies indicate a distinct outcome depending on gender. Here, we studied gender-related differences in the evolution of white matter MS-lesions (MS-WML) in early MS by using geostatistical methods. Within a 3 years observation period, a female and a male MS patient group received disease modifying drugs and underwent standardized annual brain magnetic resonance imaging, accompanied by neurological examination. MS-WML were automatically extracted and the derived binary lesion masks were subject to geostatistical analysis, yielding quantitative spatial-statistics metrics on MS-WML pattern morphology and total lesion volume (TLV). Through the MS-lesion pattern discrimination plot, the following differences were disclosed: corresponding to gender and MS-WML pattern morphology at baseline, two female subgroups (F1, F2) and two male subgroups (M1, M2) are discerned that follow a distinct MS-WML pattern evolution in space and time. F1 and M1 start with medium-level MS-WML pattern smoothness and TLV, both behave longitudinally quasi-static. By contrast, F2 and M2 start with high-level MS-WML pattern smoothness and medium-level TLV. F2 and M2 longitudinal development is characterized by strongly diminishing MS-WML pattern smoothness and TLV, i.e., continued shrinking and break-up of MS-WML. As compared to the male subgroup M2, the female subgroup F2 shows continued, increased MS-WML pattern smoothness and TLV. Data from neurological examination suggest a correlation of MS-WML pattern morphology metrics and EDSS. Our results justify detailed studies on gender-related differences.

Background: Low nurse staffing levels are associated with adverse patient outcomes from hospital care, but the causal relationship is unclear. Limited capacity to observe patients has been hypothesised as a causal mechanism. Objectives: This study determines whether or not adverse outcomes are more likely to occur after patients experience low nurse staffing levels, and whether or not missed vital signs observations mediate any relationship. Design: Retrospective longitudinal observational study. Multilevel/hierarchical mixed-effects regression models were used to explore the association between registered nurse (RN) and health-care assistant (HCA) staffing levels and outcomes, controlling for ward and patient factors. Setting and participants: A total of 138,133 admissions to 32 general adult wards of an acute hospital from 2012 to 2015. Main outcomes: Death in hospital, adverse event (death, cardiac arrest or unplanned intensive care unit admission), length of stay and missed vital signs observations. Data sources: Patient administration system, cardiac arrest database, eRoster, temporary staff bookings and the Vitalpac system (System C Healthcare Ltd, Maidstone, Kent; formerly The Learning Clinic Limited) for observations. Results: Over the first 5 days of stay, each additional hour of RN care was associated with a 3% reduction in the hazard of death [hazard ratio (HR) 0.97, 95% confidence interval (CI) 0.94 to 1.0]. Days on which the HCA staffing level fell below the mean were associated with an increased hazard of death (HR 1.04, 95% CI 1.02 to 1.07), but the hazard of death increased as cumulative staffing exposures varied from the mean in either direction. Higher levels of temporary staffing were associated with increased mortality. Adverse events and length of stay were reduced with higher RN staffing. Overall, 16% of observations were missed. Higher RN staffing was associated with fewer missed observations in high-acuity patients (incidence rate ratio 0.98, 95% CI 0.97 to 0.99), whereas the overall rate of missed observations was related to overall care hours (RN + HCA) but not to skill mix. The relationship between low RN staffing and mortality was mediated by missed observations, but other relationships between staffing and mortality were not. Changing average skill mix and staffing levels to the levels planned by the Trust, involving an increase of 0.32 RN hours per patient day (HPPD) and a similar decrease in HCA HPPD, would be associated with reduced mortality, an increase in staffing costs of £28 per patient and a saving of £0.52 per patient per hospital stay, after accounting for the value of reduced stays. Limitations: This was an observational study in a single site. Evidence of cause is not definitive. Variation in staffing could be influenced by variation in the assessed need for staff. Our economic analysis did not consider quality or length of life. Conclusions: Higher RN staffing levels are associated with lower mortality, and this study provides evidence of a causal mechanism. There may be several causal pathways and the absolute rate of missed observations cannot be used to guide staffing decisions. Increases in nursing skill mix may be cost-effective for improving patient safety. Future work: More evidence is required to validate approaches to setting staffing levels. Other aspects of missed nursing care should be explored using objective data. The implications of findings about both costs and temporary staffing need further exploration. Trial registration: This study is registered as ISRCTN17930973. Funding: This project was funded by the National Institute for Health Research (NIHR) Health Services and Delivery Research programme and will be published in full in Health Services and Delivery Research; Vol. 6, No. 38. See the NIHR Journals Library website for further project information.

ABSTRACT Classical ecology provides principles for construction and function of biological communities, but to what extent these apply to the animal-associated microbiota is just beginning to be assessed. Here, we investigated the influence of several well-known ecological principles on animal-associated microbiota by characterizing gut microbial specimens from bilaterally symmetrical animals (Bilateria) ranging from flies to whales. A rigorously vetted sample set containing 265 specimens from 64 species was assembled. Bacterial lineages were characterized by 16S rRNA gene sequencing. Previously published samples were also compared, allowing analysis of over 1,098 samples in total. A restricted number of bacterial phyla was found to account for the great majority of gut colonists. Gut microbial composition was associated with host phylogeny and diet. We identified numerous gut bacterial 16S rRNA gene sequences that diverged deeply from previously studied taxa, identifying opportunities to discover new bacterial types. The number of bacterial lineages per gut sample was positively associated with animal mass, paralleling known species-area relationships from island biogeography and implicating body size as a determinant of community stability and niche complexity. Samples from larger animals harbored greater numbers of anaerobic communities, specifying a mechanism for generating more-complex microbial environments. Predictions for species/abundance relationships from models of neutral colonization did not match the data set, pointing to alternative mechanisms such as selection of specific colonists by environmental niche. Taken together, the data suggest that niche complexity increases with gut size and that niche selection forces dominate gut community construction. IMPORTANCE The intestinal microbiome of animals is essential for health, contributing to digestion of foods, proper immune development, inhibition of pathogen colonization, and catabolism of xenobiotic compounds. How these communities assemble and persist is just beginning to be investigated. Here we interrogated a set of gut samples from a wide range of animals to investigate the roles of selection and random processes in microbial community construction. We show that the numbers of bacterial species increased with the weight of host organisms, paralleling findings from studies of island biogeography. Communities in larger organisms tended to be more anaerobic, suggesting one mechanism for niche diversification. Nonselective processes enable specific predictions for community structure, but our samples did not match the predictions of the neutral model. Thus, these findings highlight the importance of niche selection in community construction and suggest mechanisms of niche diversification.

The welfare of two genetic lines of organic layer hen pullets—H&N Super Nick (HNS) and H&N Brown Nick (HNB)—was compared during two commercial transport variants of 15 flocks of mixed-reared birds. Birds were either transported overnight (with a break in travel), or were transported direct to the layer farm (without a break in travel). Samples of feces were collected non-invasively from 25 birds of each genetic line per flock for each transport variant before transportation to evaluate baseline values of glucocorticoid metabolites, and at 0 h, 3 h, 6 h, 10 h, 24 h, 34 h, 48 h, 58 h, and 72 h after the end of transportation, to measure transportation and translocation stress. We assessed the fear toward humans with the touch test before transportation, and we checked the birds’ body condition by scoring the plumage condition and the occurrence of injuries. Body weight before and weight loss after transportation were determined, and ambient temperature was measured before, during, and after transportation. Stress investigations showed no significant differences between the transport variants (effect: −0.208; 95% confidence interval (CI): (−0.567; 0.163)). Instead, we discovered differences between the pullet lines (effect: −0.286; 95% CI: (−0.334; 0.238)). Weight loss was different between the transport variants (2.1 percentage points; 95% CI: (−2.6; −1.5)) and between the genetic lines, as HNB lost significantly less weight than HNS (0.5 percentage points; 95% CI: (0.3; 0.7)).

Aiming at iron-related T2-hypointensity, which is related to normal aging and neurodegenerative processes, we here present two practicable approaches, based on Bayesian inference, for preprocessing and statistical analysis of a complex set of structural MRI data. In particular, Markov Chain Monte Carlo methods were used to simulate posterior distributions. First, we rendered a segmentation algorithm that uses outlier detection based on model checking techniques within a Bayesian mixture model. Second, we rendered an analytical tool comprising a Bayesian regression model with smoothness priors (in the form of Gaussian Markov random fields) mitigating the necessity to smooth data prior to statistical analysis. For validation, we used simulated data and MRI data of 27 healthy controls (age: [Formula: see text]; range, [Formula: see text]). We first observed robust segmentation of both simulated T2-hypointensities and gray-matter regions known to be T2-hypointense. Second, simulated data and images of segmented T2-hypointensity were analyzed. We found not only robust identification of simulated effects but also a biologically plausible age-related increase of T2-hypointensity primarily within the dentate nucleus but also within the globus pallidus, substantia nigra, and red nucleus. Our results indicate that fully Bayesian inference can successfully be applied for preprocessing and statistical analysis of structural MRI data.