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Alaskan sled dogs develop a particular metabolic strategy during multiday submaximal exercise, allowing them to switch from intra-muscular to extra-muscular energy substrates thus postponing fatigue. Specifically, a progressively increasing stimulus for hepatic glycogenolysis and gluconeogenesis provides glucose for both fueling exercise and replenishing the depleted muscle glycogen. Moreover, recent studies have shown that with continuation of exercise sled dogs increase their insulin-sensitivity and their capacity to transport and oxidize glucose and carbohydrates rather than oxidizing fatty acids. Carnitine and acylcarnitines (AC) play an essential role as metabolic regulators in both fat and glucose metabolism; they serve as biomarkers in different species in both physiologic and pathologic conditions. We assessed the effect of multiday exercise in conditioned sled dogs on plasma short (SC), medium (MC) and long (LC) chain AC by tandem mass spectrometry (MS/MS). Our results show chain-specific modification of AC profiles during the exercise challenge: LCACs maintained a steady increase throughout exercise, some SCACs increased during the last phase of exercise and acetylcarnitine (C2) initially increased before decreasing during the later phase of exercise. We speculated that SCACs kinetics could reflect an increased protein catabolism and C2 pattern could reflect its hepatic uptake for energy-generating purposes to sustain gluconeogenesis. LCACs may be exported by muscle to avoid their accumulation to preserve glucose oxidation and insulin-sensitivity or they could be distributed by liver as energy substrates. These findings, although representing a “snapshot” of blood as a crossing point between different organs, shed further light on sled dogs metabolism that is liver-centric and more carbohydrate-dependent than fat-dependent and during prolonged submaximal exercise.

To explore what types of photographs are more helpful means to interpret natural attractions within national parks, this study focused on the relationship between the photographs with different visual characteristics and their perceived visual appeal. A photograph-based Q method was adopted. Results confirmed the visual quality of a photograph was the most important characteristic that determined its perceived attractiveness; those photographs with a high visual quality could successfully attract tourists’ attention. The subject also significantly affected the preferences of observers, suggesting an interest-dependent pattern. Using photographs of birds as examples, the participants who were interested in birds were attracted by the photographs of birds rather than those of other subjects. This study provides a better understanding of the effectiveness of photographs for communication. Findings may help researchers, communicators and national park marketers better understand and select appropriate photographs for interpretation within national parks.

BackgroundChronic hepatitis B (CHB) is endemic in the Aboriginal population of Australia's Northern Territory (NT). However, many people's hepatitis B virus (HBV) status remains unknown.Objective1. To maximise the utility of existing HBV test and vaccination data in the NT by creating a linked dataset and computerised algorithmic coding. 2. To undertake rigorous quality assurance processes to establish feasibility of using the linked dataset and computerised algorithmic coding for individual care for people living with CHB.MethodsStep 1: We used deterministic data linkage to merge information from three separate patient databases. HBV testing and vaccination data from 2008-2016 was linked and extracted for 19,314 people from 21 remote Aboriginal communities in the Top End of the NT. Step 2: A computerised algorithm was developed to allocate one of ten HBV codes to each individual. Step 3: A quality assurance process was undertaken by a clinician, using standardised processes, manually reviewing all three databases, for a subset of 5,293 Aboriginal people from five communities to check the accuracy of each allocated code.ResultsThe process of data linking individuals was highly accurate at 99.9%. The quality assurance process detected an overall error rate of 17.7% on the HBV code generated by the computerised algorithm. Errors occurred in source documentation, primarily from the historical upload of paper-based records to electronic health records. An overall HBV prevalence of 2.6% in five communities was found, which included ten cases of CHB who were previously unaware of infection and not engaged in care.ConclusionsData linkage of individuals was highly accurate. Data quality issues and poor sensitivity in the codes produced by the computerised algorithm were uncovered in the quality assurance process. By systematically, manually reviewing all available data we were able to allocate a HBV status to 91% of the study population.

BackgroundAdministrative data can be used to support research, such as in the UK Biobank. Hospital Episode Statistics (HES) are national data for England that include contain ICD-10 diagnoses for inpatient mental healthcare episodes, but the validity of these diagnoses for research purposes has not been assessed.Methods250 peoples' HES records were selected based on a HES recorded inpatient stay at the South London and Maudsley NHS Foundation Trust with a diagnosis of schizophrenia, a wider schizophrenia spectrum disorder, bipolar affective disorder or unipolar depression. A gold-standard research diagnosis was made using Clinical Records Interactive Search pseudonymised electronic patient records using, and the OPCRIT+ algorithm.ResultsPositive predictive value at the level of lifetime psychiatric disorder was 100%, and at the level of lifetime diagnosis in the four categories of schizophrenia, wider schizophrenia spectrum, bipolar or unipolar depression was 73% (68-79). Agreement varied by diagnosis, with schizophrenia having the highest PPV at 90% (80-96). Each person had an average of five psychiatric HES records. An algorithm that looked at the last recorded psychiatric diagnosis led to greatest overall agreement with the research diagnosis.DiscussionFor people who have a HES record from a psychiatric admission with a diagnosis of schizophrenia spectrum disorder, bipolar affective disorder or unipolar depression, HES records appear to be a good indicator of a mental disorder, and can provide a diagnostic category with reasonable certainty. For these diagnoses, HES records can be an effective way of ascertaining psychiatric diagnosis.

Introduction Circulating microRNAs (miRNA) are promising biomarkers for human diseases. Our study hypothesizes that circulating miRNA would reveal candidate biomarkers related to airway hyperresponsiveness (AHR) and provide biologic insights into asthma epigenetic influences. Methods Serum samples obtained at randomization for 160 children in the Childhood Asthma Management Program were profiled using a TaqMan miRNA array set. The association of the isolated miRNA with methacholine PC20 was assessed. Network and pathway analyses were performed. Functional validation of two significant miRNAs was performed in human airway smooth muscle cells (HASMs). Results Of 155 well-detected circulating miRNAs, eight were significantly associated with PC20 with the strongest association with miR-296-5p. Pathway analysis revealed miR-16-5p as a network hub, and involvement of multiple miRNAs interacting with genes in the FoxO and Hippo signaling pathways by KEGG analysis. Functional validation of two miRNA in HASM showed effects on cell growth and diameter. Conclusion Reduced circulatory miRNA expression at baseline is associated with an increase in PC20. These miRNA provide biologic insights into, and may serve as biomarkers of, asthma severity. miR-16-5p and -30d-5p regulate airway smooth muscle phenotypes critically involved in asthma pathogenesis, supporting a mechanistic link to these findings. Functional ASM phenotypes may be directly relevant to AHR.

Invasion potential should be part of the evaluation of candidate species for any species introduction. However, estimating invasion risks remains a challenging problem, particularly in complex landscapes. Certain plant traits are generally considered to increase invasive potential and there is an understanding that landscapes influence invasions dynamics, but little research has been done to explore how those drivers of invasions interact. We evaluate the relative roles of, and potential interactions between, plant invasiveness traits and landscape characteristics on invasions with a case study using a model parameterized for the potentially invasive biomass crop, Miscanthus × giganteus. Using that model we simulate invasions on 1000 real landscapes to evaluate how landscape characteristics, including both composition and spatial structure, affect invasion outcomes. We conducted replicate simulations with differing strengths of plant invasiveness traits (dispersal ability, establishment ability, population growth rate, and the ability to utilize dispersal corridors) to evaluate how the importance of landscape characteristics for predicting invasion patterns changes depending on the invader details. Analysis of simulations showed that the presence of highly suitable habitat (e.g., grasslands) is generally the strongest determinant of invasion dynamics but that there are also more subtle interactions between landscapes and invader traits. These effects can also vary between different aspects of invasion dynamics (short vs. long time scales and population size vs. spatial extent). These results illustrate that invasions are complex emergent processes with multiple drivers and effective management needs to reflect the ecology of the species of interest and the particular goals or risks for which efforts need to be optimized.

As weather patterns become more volatile and extreme, risks introduced by weather variability will become more critical to agricultural production. The availability of days suitable for field work is driven by soil temperature and moisture, both of which may be altered by climate change. We projected changes in Illinois season length, spring field workability, and summer drought risk under three different emissions scenarios (B1, A1B, and A2) down to the crop district scale. Across all scenarios, thermal time units increased in parallel with a longer frost-free season. An increase in late March and Early April field workability was consistent across scenarios, but a decline in overall April through May workable days was observed for many cases. In addition, summer drought metrics were projected to increase for most scenarios. These results highlight how the spatial and temporal variability in climate change may present unique challenges to mitigation and adaptation efforts.

At the annual Iditarod Race, Alaskan Huskies repeatedly run for up to 8 hours at 16 km/h to complete 1600 km. We previously demonstrated high rates of mitochondrial protein synthesis in Alaskan Huskies, which we suspected allowed rapid remodeling of mitochondrial proteins in response to energetic stress. The purpose of this study was to examine mitochondrial respiration in permeabilized skeletal muscle fibers of Alaskan Huskies in the offseason (Non-raced) and following the 1600 km Iditarod Sled Dog Race (Raced). We hypothesized that compared to Non-raced Huskies, raced Huskies that completed a 1600 km race would have greater mitochondrial respiratory capacities, and improvements in capacities of oxidative phosphorylation (OXPHOS) based on NADH-generating substrates as compared to fatty acids. Using high-resolution respirometry (HRR) we investigated the respiration of permeabilized muscle fibers from Alaskan Huskies. Maximum capacities were 254±26 pmol.s-1.mg-1 for OXPHOS (coupled, P) and 254±37 pmol.s-1.mg-1 for the electron transfer system (ETS; non-coupled, E). After racing respiratory capacities from NADH-linked substrates, but not fat-derived substrates increased. Finally, the OXPHOS to ETS capacity ratio (P/E) increased after racing from 0.90±0.03 to 0.97±0.02. From our previous studies and the current study, we conclude that Alaskan Huskies maintain high mitochondrial protein turnover to facilitate rapid adaptation to environmental extremes and energetic challenges.

Reconstructions of the vegetation of Europe during the Last Glacial Maximum (LGM) are an enigma. Pollen-based analyses have suggested that Europe was largely covered by steppe and tundra, and forests persisted only in small refugia. Climate-vegetation model simulations on the other hand have consistently suggested that broad areas of Europe would have been suitable for forest, even in the depths of the last glaciation. Here we reconcile models with data by demonstrating that the highly mobile groups of hunter-gatherers that inhabited Europe at the LGM could have substantially reduced forest cover through the ignition of wildfires. Similar to hunter-gatherers of the more recent past, Upper Paleolithic humans were masters of the use of fire, and preferred inhabiting semi-open landscapes to facilitate foraging, hunting and travel. Incorporating human agency into a dynamic vegetation-fire model and simulating forest cover shows that even small increases in wildfire frequency over natural background levels resulted in large changes in the forested area of Europe, in part because trees were already stressed by low atmospheric CO2 concentrations and the cold, dry, and highly variable climate. Our results suggest that the impact of humans on the glacial landscape of Europe may be one of the earliest large-scale anthropogenic modifications of the earth system.

Yield stability is fundamental to global food security in the face of climate change, and better strategies are needed for buffering crop yields against increased weather variability. Regional- scale analyses of yield stability can support robust inferences about buffering strategies for widely-grown staple crops, but have not been accomplished. We present a novel analytical approach, synthesizing 2000-2014 data on weather and soil factors to quantify their impact on county-level maize yield stability in four US states that vary widely in these factors (Illinois, Michigan, Minnesota and Pennsylvania). Yield stability is quantified as both 'downside risk' (minimum yield potential, MYP) and 'volatility' (temporal yield variability). We show that excessive heat and drought decreased mean yields and yield stability, while higher precipitation increased stability. Soil water holding capacity strongly affected yield volatility in all four states, either directly (Minnesota and Pennsylvania) or indirectly, via its effects on MYP (Illinois and Michigan). We infer that factors contributing to soil water holding capacity can help buffer maize yields against variable weather. Given that soil water holding capacity responds (within limits) to agronomic management, our analysis highlights broadly relevant management strategies for buffering crop yields against climate variability, and informs region-specific strategies.

Recently, an extensive study of 2,748 polar bears (Ursus maritimus) from across their circumpolar range was published in PLOS ONE, which used microsatellites and mitochondrial haplotypes to apparently show altered population structure and a dramatic change in directional gene flow towards the Canadian Archipelago—an area believed to be a future refugium for polar bears as their southernmost habitats decline under climate change. Although this study represents a major international collaborative effort and promised to be a baseline for future genetics work, methodological shortcomings and errors of interpretation undermine some of the study’s main conclusions. Here, we present a reanalysis of this data in which we address some of these issues, including: (1) highly unbalanced sample sizes and large amounts of systematically missing data; (2) incorrect calculation of FST and of significance levels; (3) misleading estimates of recent gene flow resulting from non-convergence of the program BayesAss. In contrast to the original findings, in our reanalysis we find six genetic clusters of polar bears worldwide: the Hudson Bay Complex, the Western and Eastern Canadian Arctic Archipelago, the Western and Eastern Polar Basin, and—importantly—we reconfirm the presence of a unique and possibly endangered cluster of bears in Norwegian Bay near Canada’s expected last sea-ice refugium. Although polar bears’ abundance, distribution, and population structure will certainly be negatively affected by ongoing—and increasingly rapid—loss of Arctic sea ice, these genetic data provide no evidence of strong directional gene flow in response to recent climate change.

BackgroundThe longitudinal birth cohort design has yielded a substantial contribution to knowledge of child health and development. The last full review in New Zealand and Australia in 2004 identified 13 studies. Since then, birth cohort designs continue to be an important tool in understanding how intrauterine, infant and childhood development affect long-term health and well-being. This updated review in a defined geographical area was conducted to better understand the factors associated with successful quality and productivity, and greater scientific and policy contribution and scope.MethodsWe adopted the preferred reporting items for systematic reviews and meta-analyses (PRISMA) approach, searching PubMed, Scopus, Cinahl, Medline, Science Direct and ProQuest between 1963 and 2013. Experts were consulted regarding further studies. Five inclusion criteria were used: (1) have longitudinally tracked a birth cohort, (2) have collected data on the child and at least one parent or caregiver (3) be based in Australia or New Zealand, (4) be empirical in design, and (5) have been published in English.Results10665 records were initially retrieved from which 23 birth cohort studies met the selection criteria. Together these studies recruited 91,196 participants, with 38,600 mothers, 14,206 fathers and 38,390 live births. Seventeen studies were located in Australia and six in New Zealand. Research questions initially focused on the perinatal period, but as studies matured, longer-term effects and outcomes were examined.ConclusionsThis review demonstrates the significant yield from this effort both in terms of scientific discovery and social policy impact. Further opportunities have been recognised with cross-study collaboration and pooling of data between established and newer studies and international studies to investigate global health determinants.

Rationale Respiratory syncytial virus (RSV) infection in preterm and newborn infants can result in severe bronchiolitis and hospitalization. The lamb lung has several key features conducive to modeling RSV infection in human infants, including susceptibility to human strains of RSV such as the A2, Long, and Memphis Strain 37 (M37). In this study, the kinetics of M37 infection was investigated in newborn lambs in order to better define clinical, viral, physiological, and immunological parameters as well as the pathology and lesions. Methods Newborn lambs were nebulized with M37 hRSV (6 mL of 1.27 x 107 FFU/mL), monitored daily for clinical responses, and respiratory tissues were collected from groups of lambs at days 1, 3, 4, 6, and 8 post-inoculation for the assessment of viral replication parameters, lesions and also cellular, immunologic and inflammatory responses. Results Lambs had increased expiratory effort (forced expiration) at days 4, 6, and 8 post-inoculation. Nasal wash lacked RSV titers at day 1, but titers were present at low levels at days 3 (peak), 4, and 8. Viral titers in bronchoalveolar lavage fluid (BALF) reached a plateau at day 3 (4.6 Log10 FFU/mL), which was maintained until day 6 (4.83 Log10 FFU/mL), and were markedly reduced or absent at day 8. Viral RNA levels (detected by RT-qPCR) in BALF were indistinguishable at days 3 (6.22 ± 0.08 Log10 M37 RNA copies/mL; mean ± se) and 4 (6.20 ± 0.16 Log10 M37 RNA copies/mL; mean ± se) and increased slightly on day 6 (7.15 ± 0.2 Log10 M37 RNA copies/mL; mean ± se). Viral antigen in lung tissue as detected by immunohistochemistry was not seen at day 1, was present at days 3 and 4 before reaching a peak by day 6, and was markedly reduced by day 8. Viral antigen was mainly present in airways (bronchi, bronchioles) at day 3 and was increasingly present in alveolar cells at days 4 and 6, with reduction at day 8. Histopathologic lesions such as bronchitis/bronchiolitis, epithelial necrosis and hyperplasia, peribronchial lymphocyte infiltration, and syncytial cells, were consistent with those described previously for lambs and infants. Conclusion This work demonstrates that M37 hRSV replication in the lower airways of newborn lambs is robust with peak replication on day 3 and sustained until day 6. These findings, along with the similarities of lamb lung to those of infants in terms of alveolar development, airway branching and epithelium, susceptibility to human RSV strains, lesion characteristics (bronchiolitis), lung size, clinical parameters, and immunity, further establish the neonatal lamb as a model with key features that mimic RSV infection in infants.

Maintaining and improving household food self-sufficiency (FSS) in mountain regions is an ongoing challenge. There are many facets to the issue, including comparatively high levels of land fragmentation, challenging terrain and transportation bottlenecks, declining labor availability due to out-migration, and low technical knowledge, among others. Using a nonparametric multivariate approach, we quantified primary associations underlying current levels of FSS in the mid-hills of Nepal. A needs assessment survey was administered to 77 households in Lungaun (Baglung District), Pang (Parbat District), and Pathlekhet (Myagdi District), with a total of 80 variables covering five performance areas; resulting data were analyzed using Classification and Regression Trees. The most parsimonious statistical model for household FSS highlighted associations with agronomic management, including yields of maize and fingermillet within a relay cropping system and adoption of improved crop cultivars. Secondary analyses of the variables retained in the first model again focused primarily on crop and livestock management. It thus appears that continued emphasis on technical agricultural improvements is warranted, independent of factors such as land holding size that, in any case, are very difficult to change through development interventions. Initiatives to increase household FSS in the mid-hills of Nepal will benefit from placing a primary focus on methods of agricultural intensification to improve crop yields and effective technology transfer to increase adoption of these methods.

Background Indigenous populations globally are disproportionately affected by chronic hepatitis B virus (HBV) infection however contemporary sero-prevalence data are often absent. In the Indigenous population of the Northern Territory (NT) of Australia the unique C4 sub-genotype of HBV universally circulates. There are no studies of the sero-prevalence, nor the impact of the vaccination program (which has a serotype mismatch compared to C4), at a population-wide level. Methods We examined all available HBV serology results obtained from the three main laboratories serving NT residents between 1991 and 2011. Data were linked with a NT government database to determine Indigenous status and the most recent test results for each individual were extracted as a cross-sectional database including 88,112 unique individuals. The primary aim was to obtain a contemporary estimate of HBsAg positivity for the NT by Indigenous status. Results Based on all tests from 2007–2011 (35,633 individuals), hepatitis B surface antigen (HBsAg) positivity was 3·40% (95%CI 3·19–3·61), being higher in Indigenous (6·08%[5·65%-6·53%]) than non-Indigenous (1·56%[1·38%-1·76%]) Australians, p

Background: Sepsis is a leading cause of death in intensive care units and is increasing in incidence. Current trials of novel therapeutic approaches for sepsis focus on 28-day mortality as the primary outcome measure, but excess mortality may extend well beyond this time period. Methods: We used relative survival analysis to examine excess mortality in a cohort of 1,028 patients admitted to a tertiary referral hospital with sepsis during 2007–2008, over the first 5 years of follow up. Expected survival was estimated using the Ederer II method, using Australian life tables as the reference population. Cumulative and interval specific relative survival were estimated by age group, sex, sepsis severity and Indigenous status. Results : Patients were followed for a median of 4.5 years (range 0–5.2). Of the 1028 patients, the mean age was 46.9 years, 52% were male, 228 (22.2%) had severe sepsis and 218 (21%) died during the follow up period. Mortality based on cumulative relative survival exceeded that of the reference population for the first 2 years post admission in the whole cohort and for the first 3 years in the subgroup with severe sepsis. Independent predictors of mortality over the whole follow up period were male sex, Indigenous Australian ethnicity, older age, higher Charlson Comorbidity Index, and sepsis-related organ dysfunction at presentation. Conclusions: The mortality rate of patients hospitalised with sepsis exceeds that of the general population until 2 years post admission. Efforts to improve outcomes from sepsis should examine longer term outcomes than the traditional primary endpoints of 28-day and 90-day mortality.

Endurance exercise relies on transsarcolemmal flux of substrates in order to avoid depletion of intramuscular reserves. Previous studies of endurance trained sled dogs have shown a remarkable capacity of these dogs to adapt rapidly to endurance exercise by decreasing the utilization of intramuscular reserves. The current study tested the hypothesis that the dogs' glycogen-sparing phenotype is due to increased sarcolemmal transport of glucose and fatty acids. Basal and exercise-induced transport of glucose and fatty acids into sarcolemmal vesicles was evaluated in racing sled dogs prior to and after 7 months of exercise conditioning. Sarcolemmal substrate transport capacity was measured using sarcolemmal vesicles and radiolabelled substrates, and transporter abundance was measured using Western blot quantification in whole muscle homogenates and the sarcolemmal vesicle preparations. Conditioning resulted in increased basal and exercise-induced transport of both glucose and palmitate. Neither acute exercise nor conditioning resulted in changes in muscle content of GLUT4 or FAT/CD36, but conditioning did result in decreased abundance of both transporters in the sarcolemmal vesicles used for the basal transport assays, and this decrease was further amplified in the vesicles used for the exercise-induced transport assays. These results demonstrate conditioning-induced increases in sarcolemmal transport of oxidizable substrates, as well as increased gain of exercise-induced sarcolemmal transport of these substrates. These results further indicate that increased sarcolemmal transport of oxidizable substrates may be due to either an increased intrinsic capacity of the existing transporters or to a different population of transporters from those investigated.

The seed stage is often critical in determining the regeneration success of plants. Seeds must survive an array of seed predators and pathogens and germinate under conditions favourable for seedling establishment. To maximise recruitment success plants protect seeds using a diverse set of chemical and physical defences. However, the relationship between these defence classes, and their association with other life history traits, is not well understood. Data on seed coat thickness and fracture resistance, and the abundance and diversity of potential defensive compounds were collected for 10 tree species of Macaranga from Borneo. The data were used to test whether there is a trade-off in physical versus chemical defence investment, and to determine how investment varies with seed mass, and light requirement for regeneration. Across species there was no correlation between seed coat thickness and abundance of potential defensive compounds, indicating the absence of a direct trade-off between defence classes. While chemical defences were not correlated to other traits, physical defences were positively correlated with light requirement for regeneration. For a subset of five Macaranga species we evaluated the relative investment in chemical and physical defence to seed persistence in the soil, measured as the time to half initial seed viability (seed half-life). Half-life was negatively related to the ratio of potential defensive compound abundance to seed coat thickness, suggesting that species with long persistence invested in physical defence more than stored chemical defences. These results indicate that investment in seed defences are associated with species' light requirements for regeneration, rather than scaling positively with seed mass. Furthermore, chemical defences, although highly variable among species, do not appear to be critical to long term persistence of Macaranga seeds, and may be important in defending seeds from natural enemies distinct from those found in the soil.

Renal medullary interstitial cells (RMIC) are specialized fibroblast-like cells that exert important functions in maintaining body fluid homeostasis and systemic blood pressure. Here, we generated a RMIC specific tenascin-C promoter driven inducible CreER2 knockin mouse line with an EGFP reporter. Similar as endogenous tenascin-C expression, the reporter EGFP expression in the tenascin-C-CreER2(+/-) mice was observed in the inner medulla of the kidney, and co-localized with COX2 but not with AQP2 or AQP1, suggesting selective expression in RMICs. After recombination (tenascin-C-CreER2(+/-)/ROSA26-lacZ(+/-) mice + tamoxifen), β-gal activity was restricted to the cells in the inner medulla of the kidney, and didn't co-localize with AQP2, consistent with selective Cre recombinase activity in RMICs. Cre activity was not obvious in other major organs or without tamoxifen treatment. This inducible RMIC specific Cre mouse line should therefore provide a novel tool to manipulate genes of interest in RMICs.

In interpersonal communication, the listener can often see as well as hear the speaker. Visual stimuli can subtly change a listener’s auditory perception, as in the McGurk illusion, in which perception of a phoneme’s auditory identity is changed by a concurrent video of a mouth articulating a different phoneme. Studies have yet to link visual influences on the neural representation of language with subjective language perception. Here we show that vision influences the electrophysiological representation of phonemes in human auditory cortex prior to the presentation of the auditory stimulus. We used the McGurk effect to dissociate the subjective perception of phonemes from the auditory stimuli. With this paradigm we demonstrate that neural representations in auditory cortex are more closely correlated with the visual stimuli of mouth articulation, which drive the illusory subjective auditory perception, than the actual auditory stimuli. Additionally, information about visual and auditory stimuli transfer in the caudal–rostral direction along the superior temporal gyrus during phoneme perception as would be expected of visual information flowing from the occipital cortex into the ventral auditory processing stream. These results show that visual stimuli influence the neural representation in auditory cortex early in sensory processing and may override the subjective auditory perceptions normally generated by auditory stimuli. These findings depict a marked influence of vision on the neural processing of audition in tertiary auditory cortex and suggest a mechanistic underpinning for the McGurk effect.

Background MicroRNAs are key transcriptional and network regulators previously associated with asthma susceptibility. However, their role in relation to asthma severity has not been delineated. Objective We hypothesized that circulating microRNAs could serve as biomarkers of changes in lung function in asthma patients. Methods We isolated microRNAs from serum samples obtained at randomization for 160 participants of the Childhood Asthma Management Program. Using a TaqMan microRNA array containing 754 microRNA primers, we tested for the presence of known asthma microRNAs, and assessed the association of the individual microRNAs with lung function as measured by FEV1/FVC, FEV1% and FVC%. We further tested the subset of FEV1/FVC microRNAs for sex-specific and lung developmental associations. Results Of the 108 well-detected circulating microRNAs, 74 (68.5%) had previously been linked to asthma susceptibility. We found 22 (20.3%), 4 (3.7%) and 8 (7.4%) microRNAs to be associated with FEV1/FVC, FEV1% and FVC%, respectively. 8 (of 22) FEV1/FVC, 3 (of 4) FEV1% and 1 (of 8) FVC% microRNAs had functionally validated target genes that have been linked via genome wide association studies to asthma and FEV1 change. Among the 22 FEV1/FVC microRNAs, 9 (40.9%) remain associated with FEV1/FVC in boys alone in a sex-stratified analysis (compared with 3 FEV1/FVC microRNAs in girls alone), 7 (31.8%) were associated with fetal lung development, and 3 (13.6%) in both. Ontology analyses revealed enrichment for pathways integral to asthma, including PPAR signaling, G-protein coupled signaling, actin and myosin binding, and respiratory system development. Conclusions Circulating microRNAs reflect asthma biology and are associated with lung function differences in asthmatics. They may represent biomarkers of asthma severity.

Background and Methods Mice are used for modelling the biology of many human diseases, including colorectal cancer (CRC). Mouse models recapitulate many aspects of human disease and are invaluable tools for studying the biology, treatment and prevention of CRC. Unlike humans, many mouse models develop lesions primarily in the small intestine, which necessitates removal and examination of this organ in order to evaluate treatment efficacy. Commonly, the small intestine is visually examined for gross lesions and then selectively embedded in paraffin blocks for further microscopic analysis. Unfortunately, this method suffers from inherent bias toward counting large lesions and simultaneously missing smaller lesions. Even more, this method leaves no permanent record of diagnosed and measured lesions. We evaluated inter-observer variability in a mouse model of CRC using visual examination, and directly compared the visual, gross examination with a histologic analytic method using digital slides of hematoxylin and eosin stained tissue sections. Results Using visual examination, there was a high degree of inter-observer variability. As this method does not provide a permanent record of measurements, there is no capability to arbitrate between differing observations. In contrast, histologic analysis allowed for the creation of a permanent record of lesion measurements taken. When compared directly, histologic analysis of annotated digital images has significantly improved accuracy. Using this method we were able to distinguish mutant mice from wild type littermates even at a very young age. With gross visual examination, this distinction was not possible. Conclusion Histologic analysis of digital images of murine intestinal tissue provides a vital improvement over the commonly used visual, gross examination method. Unlike visual gross examination, histologic analysis is not biased by the size of intestinal adenoma, misdiagnosis of another lesion type, or presence of a Peyer’s patch. It also provides accountability in the form of a permanent record of lesions counted. Histologic analysis using digital slides represents a critical improvement over the current, widely used method of visual gross examination and should be considered for future studies using mouse models of CRC.

Balancing productivity, profitability, and environmental health is a key challenge for agricultural sustainability. Most crop production systems in the United States are characterized by low species and management diversity, high use of fossil energy and agrichemicals, and large negative impacts on the environment. We hypothesized that cropping system diversification would promote ecosystem services that would supplement, and eventually displace, synthetic external inputs used to maintain crop productivity. To test this, we conducted a field study from 2003–2011 in Iowa that included three contrasting systems varying in length of crop sequence and inputs. We compared a conventionally managed 2-yr rotation (maize-soybean) that received fertilizers and herbicides at rates comparable to those used on nearby farms with two more diverse cropping systems: a 3-yr rotation (maize-soybean-small grain + red clover) and a 4-yr rotation (maize-soybean-small grain + alfalfa-alfalfa) managed with lower synthetic N fertilizer and herbicide inputs and periodic applications of cattle manure. Grain yields, mass of harvested products, and profit in the more diverse systems were similar to, or greater than, those in the conventional system, despite reductions of agrichemical inputs. Weeds were suppressed effectively in all systems, but freshwater toxicity of the more diverse systems was two orders of magnitude lower than in the conventional system. Results of our study indicate that more diverse cropping systems can use small amounts of synthetic agrichemical inputs as powerful tools with which to tune, rather than drive, agroecosystem performance, while meeting or exceeding the performance of less diverse systems.

Granulosa cell tumors (GCTs) are the most common ovarian estrogen producing tumors, leading to symptoms of excessive estrogen such as endometrial hyperplasia and endometrial adenocarcinoma. These tumors have malignant potential and often recur. The etiology of GCT is unknown. TGFα is a potent mitogen for many different cells. However, its function in GCT initiation, progression and metastasis has not been determined. The present study aims to determine whether TGFα plays a role in the growth of GCT cells. KGN cells, which are derived from an invasive GCT and have many features of normal granulosa cells, were used as the cellular model. Immunohistochemistry, Western blot and RT-PCR results showed that the ErbB family of receptors is expressed in human GCT tissues and GCT cell lines. RT-PCR results also indicated that TGFα and EGF are expressed in the human granulosa cells and the GCT cell lines, suggesting that TGFα might regulate GCT cell function in an autocrine/paracrine manner. TGFα stimulated KGN cell DNA synthesis, cell proliferation, cell viability, cell cycle progression, and cell migration. TGFα rapidly activated EGFR/PI3K/Akt and mTOR pathways, as indicated by rapid phosphorylation of Akt, TSC2, Rictor, mTOR, P70S6K and S6 proteins following TGFα treatment. TGFα also rapidly activated the EGFR/MEK/ERK pathway, and P38 MAPK pathways, as indicated by the rapid phosphorylation of EGFR, MEK, ERK1/2, P38, and CREB after TGFα treatment. Whereas TGFα triggered a transient activation of Akt, it induced a sustained activation of ERK1/2 in KGN cells. Long-term treatment of KGN cells with TGFα resulted in a significant increase in cyclin D2 and a decrease in p27/Kip1, two critical regulators of granulosa cell proliferation and granulosa cell tumorigenesis. In conclusion, TGFα, via multiple signaling pathways, regulates KGN cell proliferation and migration and may play an important role in the growth and metastasis of GCTs.

Both endothelial and immune dysfunction contribute to the high mortality rate in human sepsis, but the underlying mechanisms are unclear. In response to infection, interferon-c activates indoleamine 2,3-dioxygenase (IDO) which metabolizes the essential amino acid tryptophan to the toxic metabolite kynurenine. IDO can be expressed in endothelial cells, hepatocytes and mononuclear leukocytes, all of which contribute to sepsis pathophysiology. Increased IDO activity (measured by the kynurenine to tryptophan [KT] ratio in plasma) causes T-cell apoptosis, vasodilation and nitric oxide synthase inhibition. We hypothesized that IDO activity in sepsis would be related to plasma interferon-c, interleukin-10, T cell lymphopenia and impairment of microvascular reactivity, a measure of endothelial nitric oxide bioavailability. In an observational cohort study of 80 sepsis patients (50 severe and 30 non-severe) and 40 hospital controls, we determined the relationship between IDO activity (plasma KT ratio) and selected plasma cytokines, sepsis severity, nitric oxide-dependent microvascular reactivity and lymphocyte subsets in sepsis. Plasma amino acids were measured by high performance liquid chromatography and microvascular reactivity by peripheral arterial tonometry. The plasma KT ratio was increased in sepsis (median 141 [IQR 64–235]) compared to controls (36 [28–52]); p,0.0001), and correlated with plasma interferon-c and interleukin-10, and inversely with total lymphocyte count, CD8+ and CD4+ T-lymphocytes, systolic blood pressure and microvascular reactivity. In response to treatment of severe sepsis, the median KT ratio decreased from 162 [IQR 100–286] on day 0 to 89 [65–139] by day 7; p = 0.0006) and this decrease in KT ratio correlated with a decrease in the Sequential Organ Failure Assessment score (p,0.0001). IDO-mediated tryptophan catabolism is associated with dysregulated immune responses and impaired microvascular reactivity in sepsis and may link these two fundamental processes in sepsis pathophysiology.

The white-tailed deer (Odocoileus virginianus) represents one of the most successful and widely distributed large mammal species within North America, yet very little nucleotide sequence information is available. We utilized massively parallel pyrosequencing of a reduced representation library (RRL) and a random shotgun library (RSL) to generate a complete mitochondrial genome sequence and identify a large number of putative single nucleotide polymorphisms (SNPs) distributed throughout the white-tailed deer nuclear and mitochondrial genomes. A SNP validation study designed to test specific classes of putative SNPs provides evidence for as many as 10,476 genome-wide SNPs in the current dataset. Based on cytogenetic evidence for homology between cow (Bos taurus) and white-tailed deer chromosomes, we demonstrate that a divergent genome may be used for estimating the relative distribution and density of de novo sequence contigs as well as putative SNPs for species without draft genome assemblies. Our approach demonstrates that bioinformatic tools developed for model or agriculturally important species may be leveraged to support next-generation research programs for species of biological, ecological and evolutionary importance. We also provide a functional annotation analysis for the de novo sequence contigs assembled from white-tailed deer pyrosequencing reads, a mitochondrial phylogeny involving 13,722 nucleotide positions for 10 unique species of Cervidae, and a median joining haplotype network as a putative representation of mitochondrial evolution in O. virginianus. The results of this study are expected to provide a detailed template enabling genome-wide sequence-based studies of threatened, endangered or conservationally important non-model organisms.

RNA sequencing (RNA-seq) enables characterization and quantification of individual transcriptomes as well as detection of patterns of allelic expression and alternative splicing. Current RNA-seq protocols depend on high-throughput short-read sequencing of cDNA. However, as ongoing advances are rapidly yielding increasing read lengths, a technical hurdle remains in identifying the degree to which differences in read length influence various transcriptome analyses. In this study, we generated two paired-end RNA-seq datasets of differing read lengths (2x75 bp and 2x262 bp) for lymphoblastoid cell line GM12878 and compared the effect of read length on transcriptome analyses, including read-mapping performance, gene and transcript quantification, and detection of allele-specific expression (ASE) and allele-specific alternative splicing (ASAS) patterns. Our results indicate that, while the current long-read protocol is considerably more expensive than short-read sequencing, there are important benefits that can only be achieved with longer read length, including lower mapping bias and reduced ambiguity in assigning reads to genomic elements, such as mRNA transcript. We show that these benefits ultimately lead to improved detection of cis-acting regulatory and splicing variation effects within individuals., 29 pages, 8 figures, 11 supplementary figures

Studies utilizing general learning and memory tasks have suggested the importance of neocortical structural plasticity for memory consolidation. However, these learning tasks typically result in learning of multiple different tasks over several days of training, making it difficult to determine the synaptic time course mediating each learning event. The current study used trace-eyeblink conditioning to determine the time course for neocortical spine modification during learning. With eyeblink conditioning, subjects are presented with a neutral, conditioned stimulus (CS) paired with a salient, unconditioned stimulus (US) to elicit an unconditioned response (UR). With multiple CS-US pairings, subjects learn to associate the CS with the US and exhibit a conditioned response (CR) when presented with the CS. Trace conditioning is when there is a stimulus free interval between the CS and the US. Utilizing trace-eyeblink conditioning with whisker stimulation as the CS (whisker-trace-eyeblink: WTEB), previous findings have shown that primary somatosensory (barrel) cortex is required for both acquisition and retention of the trace-association. Additionally, prior findings demonstrated that WTEB acquisition results in an expansion of the cytochrome oxidase whisker representation and synaptic modification in layer IV of barrel cortex. To further explore these findings and determine the time course for neocortical learning-induced spine modification, the present study utilized WTEB conditioning to examine Golgi-Cox stained neurons in layer IV of barrel cortex. Findings from this study demonstrated a training-dependent spine proliferation in layer IV of barrel cortex during trace associative learning. Furthermore, findings from this study showing that filopodia-like spines exhibited a similar pattern to the overall spine density further suggests that reorganization of synaptic contacts set the foundation for learning-induced neocortical modifications through the different neocortical layers.

Here we describe Casuarina virus (CASV), a new virus in the family Mesoniviridae. This is the first report of a mesonivirus in Australia, which extends the geographical range of this virus family to 3 continents. The virus was isolated in 2010 from Coquillettidia xanthogaster mosquitoes during surveillance in the suburbs of Darwin, the capital of the Northern Territory. Cryo-electron microscopy of the CASV virions revealed spherical particles of 65 nm in size with large club-shaped projections of approximately 15 nm in length. The new virus was most closely related to Alphamesonivirus 1, the only currently recognized species in the family. In 2013 a further 5 putative new mesonivirus species were described: Hana, Méno, Nsé, Moumo and Dak Nong viruses. The evolutionary distance between CASV and two of its closest relatives, Cavally and Hana viruses (Jones-Taylor-Thornton distance of 0.151 and 0.224, respectively), along with its isolation from a different genus of mosquitoes captured on a separate continent indicate that CASV is a new species.

Free-ranging marine predators rarely search for prey along straight lines because dynamic ocean processes usually require complex search strategies. If linear movement patterns occur they are usually associated with travelling events or migratory behaviour. However, recent fine scale tracking of flying seabirds has revealed straight-line movements while birds followed fishing vessels. Unlike flying seabirds, penguins are not known to target and follow fishing vessels. Yet yellow-eyed penguins from New Zealand often exhibit directed movement patterns while searching for prey at the seafloor, a behaviour that seems to contradict common movement ecology theories. While deploying GPS dive loggers on yellow-eyed penguins from the Otago Peninsula we found that the birds frequently followed straight lines for several kilometres with little horizontal deviation. In several cases individuals swam up and down the same line, while some of the lines were followed by more than one individual. Using a remote operated vehicle (ROV) we found a highly visible furrow on the seafloor most likely caused by an otter board of a demersal fish trawl, which ran in a straight line exactly matching the trajectory of a recent line identified from penguin tracks. We noted high abundances of benthic scavengers associated with fisheries-related bottom disturbance. While our data demonstrate the acute way-finding capabilities of benthic foraging yellow-eyed penguins, they also highlight how hidden cascading effects of coastal fisheries may alter behaviour and potentially even population dynamics of marine predators, an often overlooked fact in the examination of fisheries' impacts.

Retinitis pigmentosa (RP) and Leber congenital amaurosis (LCA) are major causes of blindness. They result from mutations in many genes which has long hampered comprehensive genetic analysis. Recently, targeted next-generation sequencing (NGS) has proven useful to overcome this limitation. To uncover "hidden mutations" such as copy number variations (CNVs) and mutations in non-coding regions, we extended the use of NGS data by quantitative readout for the exons of 55 RP and LCA genes in 126 patients, and by including non-coding 5' exons. We detected several causative CNVs which were key to the diagnosis in hitherto unsolved constellations, e.g. hemizygous point mutations in consanguineous families, and CNVs complemented apparently monoallelic recessive alleles. Mutations of non-coding exon 1 of EYS revealed its contribution to disease. In view of the high carrier frequency for retinal disease gene mutations in the general population, we considered the overall variant load in each patient to assess if a mutation was causative or reflected accidental carriership in patients with mutations in several genes or with single recessive alleles. For example, truncating mutations in RP1, a gene implicated in both recessive and dominant RP, were causative in biallelic constellations, unrelated to disease when heterozygous on a biallelic mutation background of another gene, or even non-pathogenic if close to the C-terminus. Patients with mutations in several loci were common, but without evidence for di- or oligogenic inheritance. Although the number of targeted genes was low compared to previous studies, the mutation detection rate was highest (70%) which likely results from completeness and depth of coverage, and quantitative data analysis. CNV analysis should routinely be applied in targeted NGS, and mutations in non-coding exons give reason to systematically include 5'-UTRs in disease gene or exome panels. Consideration of all variants is indispensable because even truncating mutations may be misleading.

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that is characterized by defective immune tolerance combined with immune cell hyperactivity resulting in the production of pathogenic autoantibodies. Previous gene expression studies employing whole blood or peripheral blood mononuclear cells (PBMC) have demonstrated that a majority of patients with active disease have increased expression of type I interferon (IFN) inducible transcripts known as the IFN signature. The goal of the current study was to assess the gene expression profiles of isolated leukocyte subsets obtained from SLE patients. Subsets including CD19(+) B lymphocytes, CD3(+)CD4(+) T lymphocytes and CD33(+) myeloid cells were simultaneously sorted from PBMC. The SLE transcriptomes were assessed for differentially expressed genes as compared to healthy controls. SLE CD33(+) myeloid cells exhibited the greatest number of differentially expressed genes at 208 transcripts, SLE B cells expressed 174 transcripts and SLE CD3(+)CD4(+) T cells expressed 92 transcripts. Only 4.4% (21) of the 474 total transcripts, many associated with the IFN signature, were shared by all three subsets. Transcriptional profiles translated into increased protein expression for CD38, CD63, CD107a and CD169. Moreover, these studies demonstrated that both SLE lymphoid and myeloid subsets expressed elevated transcripts for cytosolic RNA and DNA sensors and downstream effectors mediating IFN and cytokine production. Prolonged upregulation of nucleic acid sensing pathways could modulate immune effector functions and initiate or contribute to the systemic inflammation observed in SLE.

Recent reports of a novel group of flaviviruses that replicate only in mosquitoes and appear to spread through insect populations via vertical transmission have emerged from around the globe. To date, there is no information on the presence or prevalence of these insect-specific flaviviruses (ISFs) in Australian mosquito species. To assess whether such viruses occur locally, we used reverse transcription-polymerase chain reaction (RT-PCR) and flavivirus universal primers that are specific to the NS5 gene to detect these viruses in mosquito pools collected from the Northern Territory. Of 94 pools of mosquitoes, 13 were RT-PCR positive, and of these, 6 flavivirus isolates were obtained by inoculation of mosquito cell culture. Sequence analysis of the NS5 gene revealed that these isolates are genetically and phylogenetically similar to ISFs reported from other parts of the world. The entire coding region of one isolate (designated 56) was sequenced and shown to have approximately 63.7% nucleotide identity and 66.6% amino acid identity with its closest known relative (Nakiwogo virus) indicating that the prototype Australian ISF represents a new species. All isolates were obtained from Coquillettidia xanthogaster mosquitoes. The new virus is tentatively named Palm Creek virus (PCV) after its place of isolation. We also demonstrated that prior infection of cultured mosquito cells with PCV suppressed subsequent replication of the medically significant West Nile and Murray Valley encephalitis viruses by 10-43 fold (1 to 1.63 log) at 48 hr post-infection, suggesting that superinfection exclusion can occur between ISFs and vertebrate-infecting flaviviruses despite their high level of genetic diversity. We also generated several monoclonal antibodies (mAbs) that are specific to the NS1 protein of PCV, and these represent the first ISF-specific mAbs reported to date. Refereed/Peer-reviewed

Background: Plasma concentrations of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide synthase, are raised in patients with chronic vascular disease, causing increased cardiovascular risk and endothelial dysfunction, but the role of ADMA in acute inflammatory states is less well defined. Methods and Results: In a prospective longitudinal study in 67 patients with acute sepsis and 31 controls, digital microvascular reactivity was measured by peripheral arterial tonometry and blood was collected at baseline and 2–4 days later. Plasma ADMA and L-arginine concentrations were determined by high performance liquid chromatography. Baseline plasma L-arginine: ADMA ratio was significantly lower in sepsis patients (median [IQR] 63 [45–103]) than in hospital controls (143 [123–166], p,0.0001) and correlated with microvascular reactivity (r = 0.34, R2 = 0.12, p = 0.02). Baseline plasma ADMA was independently associated with 28-day mortality (Odds ratio [95% CI] for death in those in the highest quartile ($0.66 mmol/L) = 20.8 [2.2–195.0], p = 0.008), and was independently correlated with severity of organ failure. Increase in ADMA over time correlated with increase in organ failure and decrease in microvascular reactivity. Conclusions: Impaired endothelial and microvascular function due to decreased endothelial NO bioavailability is a potential mechanism linking increased plasma ADMA with organ failure and death in sepsis.