CoESPRIT: A Library-Based Construct Screening Method for Identification and Expression of Soluble Protein Complexes

Structural and biophysical studies of protein complexes require multi-milligram quantities of soluble material. Subunits are often unstable when expressed separately so co-expression strategies are commonly employed since in vivo complex formation can provide stabilising effects. Defining constructs for subunit co-expression experiments is difficult if the proteins are poorly understood. Even more problematic is when subunit polypeptide chains co-fold since individually they do not have predictable domains. We have developed CoESPRIT, a modified version of the ESPRIT random library construct screen used previously on single proteins, to express soluble protein complexes. A random library of target constructs is screened against a fixed bait protein to identify stable complexes. In a proof-of-principle study, C-terminal fragments of the influenza polymerase PB2 subunit containing folded domains were isolated using importin alpha as bait. Separately, a C-terminal fragment of the PB1 subunit was used as bait to trap N-terminal fragments of PB2 resulting in co-folded complexes. Subsequent expression of the target protein without the bait indicates whether the target is independently stable, or co-folds with its partner. This highly automated method provides an efficient strategy for obtaining recombinant protein complexes at yields compatible with structural, biophysical and functional studies

All Thresholds Barred: Direct Estimation of Call Density in Bioacoustic Data

Passive acoustic monitoring (PAM) studies generate thousands of hours of audio, which may be used to monitor specific animal populations, conduct broad biodiversity surveys, detect threats such as poachers, and more. Machine learning classifiers for species identification are increasingly being used to process the vast amount of audio generated by bioacoustic surveys, expediting analysis and increasing the utility of PAM as a management tool. In common practice, a threshold is applied to classifier output scores, and scores above the threshold are aggregated into a detection count. The choice of threshold produces biased counts of vocalizations, which are subject to false positive/negative rates that may vary across subsets of the dataset. In this work, we advocate for directly estimating call density: The proportion of detection windows containing the target vocalization, regardless of classifier score. Our approach targets a desirable ecological estimator and provides a more rigorous grounding for identifying the core problems caused by distribution shifts -- when the defining characteristics of the data distribution change -- and designing strategies to mitigate them. We propose a validation scheme for estimating call density in a body of data and obtain, through Bayesian reasoning, probability distributions of confidence scores for both the positive and negative classes. We use these distributions to predict site-level densities, which may be subject to distribution shifts. We test our proposed methods on a real-world study of Hawaiian birds and provide simulation results leveraging existing fully annotated datasets, demonstrating robustness to variations in call density and classifier model quality.Comment: 14 pages, 6 figures, 3 tables; submitted to Frontiers in Bird Science; Our Hawaiian PAM dataset and classifier scores, as well as annotation information for the three study species, can be found on Zenodo at https://doi.org/10.5281/zenodo.10581530. The fully annotated Powdermill dataset assembled by Chronister et al. that was used in this study is available at https://doi.org/10.1002/ecy.332

Widespread gas hydrate instability on the upper U.S. Beaufort margin

Author Posting. © American Geophysical Union, 2014. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Journal of Geophysical Research: Solid Earth 119 (2014): 8594–8609, doi:10.1002/2014JB011290.The most climate-sensitive methane hydrate deposits occur on upper continental slopes at depths close to the minimum pressure and maximum temperature for gas hydrate stability. At these water depths, small perturbations in intermediate ocean water temperatures can lead to gas hydrate dissociation. The Arctic Ocean has experienced more dramatic warming than lower latitudes, but observational data have not been used to study the interplay between upper slope gas hydrates and warming ocean waters. Here we use (a) legacy seismic data that constrain upper slope gas hydrate distributions on the U.S. Beaufort Sea margin, (b) Alaskan North Slope borehole data and offshore thermal gradients determined from gas hydrate stability zone thickness to infer regional heat flow, and (c) 1088 direct measurements to characterize multidecadal intermediate ocean warming in the U.S. Beaufort Sea. Combining these data with a three-dimensional thermal model shows that the observed gas hydrate stability zone is too deep by 100 to 250 m. The disparity can be partially attributed to several processes, but the most important is the reequilibration (thinning) of gas hydrates in response to significant (~0.5°C at 2σ certainty) warming of intermediate ocean temperatures over 39 years in a depth range that brackets the upper slope extent of the gas hydrate stability zone. Even in the absence of additional ocean warming, 0.44 to 2.2 Gt of methane could be released from reequilibrating gas hydrates into the sediments underlying an area of ~5–7.5 × 103 km2 on the U.S. Beaufort Sea upper slope during the next century.This work was supported by the U.S. Department of Energy (DOE), grant DE-FE0010180 to SMU and a USGS-DOE interagency agreement DE-FE0005806.2015-06-0

The Wide Field Spectrograph (WiFeS)

This paper describes the Wide Field Spectrograph (WiFeS) under construction at the Research School of Astronomy and Astrophysics (RSAA) of the Australian National University (ANU) for the ANU 2.3m telescope at the Siding Spring Observatory. WiFeS is a powerful integral field, double-beam, concentric, image-slicing spectrograph designed to deliver excellent thoughput, wavelength stability, spectrophotometric performance and superb image quality along with wide spectral coverage throughout the 320-950 nm wavelength region. It provides a 25x38 arcsec. field with 0.5 arcsec. sampling along each of twenty five 38X1 arcsec slitlets. The output format is optimized to match the 4096x4096 pixel CCD detectors in each of two cameras individually optimized for the blue and the red ends of the spectrum, respectively. A process of "interleaved nod-and-shuffle" will be applied to permit quantum noise-limited sky subtraction. Using VPH gratings, spectral resolutions of 3000 and 7000 are provided. The full spectral range is covered in a single exposure at R=3000, and in two exposures in the R=7000 mode. The use of transmissive coated optics, VPH gratings and optimized mirror coatings ensures a throughput (including telescope atmosphere and detector) > 30% over a wide spectral range. The concentric image-slicer design ensures an excellent and uniform image quality across the full field. To maximize scientific return, the whole instrument is configured for remote observing, pipeline data reduction, and the accumulation of calibration image libraries.Comment: Accepted for publication in Astrophysics & Space Science, 16 pages, 14 figure

Musculoskeletal injury epidemiology in law enforcement and firefighter recruits during physical training: a systematic review

Objectives: Report the injury epidemiology of law enforcement and firefighter recruits. Design: A systematic epidemiological review following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses 2020 guidelines was completed. Data sources: Five online databases were searched from database inception to 5 May 2021. Eligibility criteria for selecting studies: Prospective and retrospective studies that reported data on musculoskeletal injuries sustained by law enforcement or firefighter recruits were included. We reported on all components of injury where data were available. All injury incidence rates were calculated as per 1000 training days (Poisson 95% CI) to allow comparisons between studies. Study quality was assessed using the Joanna Briggs Institute Quality Assessment Checklist for Prevalence Studies. Results: No studies reporting firefighter recruits were identified. Eight published studies that reported on injuries to law enforcement recruits were identified. The studies were all low quality, and the credibility of the evidence was assessed as very low. Seven studies reported medical attention injuries, and one study reported the number of medical withdrawals from a recruit training programme. The prevalence of law enforcement recruits with medical attention injuries ranged from 13.7% to 24.5%. The overall medical attention injury incidence rate for law enforcement recruits ranged from 1.67 injuries per 1000 training days (Poisson 95% CI 1.00 to 2.34 injuries per 1000 training days) to 4.24 injuries per 1000 training days (Poisson 95% CI 2.97 to 5.51 injuries per 1000 training days). Conclusion: This review reported the prevalence and incidence rates for musculoskeletal injuries in law enforcement officers. However, the credibility of the evidence is very low

Evidence for Placoderms from the Mid-Palaeozoic Sandon Beds of North-western New South Wales, Australia

Armoured jawed fishes known as placoderms are a well-documented group with a fossil record spanning the Silurian to end-Devonian. They have a global distribution and a marked diversity within Devonian deposits of Australia. Despite their notable Gondwanan fossil record, new material is occasionally identified and can present important stratigraphic information for otherwise under-explored deposits. A unique find from the so-called Sandon beds is presented here and expands the record of placoderms from New South Wales. This specimen presents insight into a previously unknown macrofossil record from the deposit and suggests a more Devonian age for the unit, rather than the previously suggested Carboniferous date. We also summarise the macrovertebrate record of Devonian placoderms from Australia, highlighting and discussing changes in their Gondwanan taxonomic diversity across the time period

Genetic structure along an elevational gradient in Hawaiian honeycreepers reveals contrasting evolutionary responses to avian malaria

<p>Abstract</p> <p>Background</p> <p>The Hawaiian honeycreepers (Drepanidinae) are one of the best-known examples of an adaptive radiation, but their persistence today is threatened by the introduction of exotic pathogens and their vector, the mosquito <it>Culex quinquefasciatus</it>. Historically, species such as the amakihi (<it>Hemignathus virens</it>), the apapane (<it>Himatione sanguinea</it>), and the iiwi (<it>Vestiaria coccinea</it>) were found from the coastal lowlands to the high elevation forests, but by the late 1800's they had become extremely rare in habitats below 900 m. Recently, however, populations of amakihi and apapane have been observed in low elevation habitats. We used twelve polymorphic microsatellite loci to investigate patterns of genetic structure, and to infer responses of these species to introduced avian malaria along an elevational gradient on the eastern flanks of Mauna Loa and Kilauea volcanoes on the island of Hawaii.</p> <p>Results</p> <p>Our results indicate that amakihi have genetically distinct, spatially structured populations that correspond with altitude. We detected very few apapane and no iiwi in low-elevation habitats, and genetic results reveal only minimal differentiation between populations at different altitudes in either of these species.</p> <p>Conclusion</p> <p>Our results suggest that amakihi populations in low elevation habitats have not been recolonized by individuals from mid or high elevation refuges. After generations of strong selection for pathogen resistance, these populations have rebounded and amakihi have become common in regions in which they were previously rare or absent.</p