Sampling Artifacts Research Articles

Illusion of flight? Absence, evidence and the age of winged insects

Abstract The earliest fossils of winged insects (Pterygota) are mid-Carboniferous (latest Mississippian, 328–324 Mya), but estimates of their age based on fossil-calibrated molecular phylogenetic studies place their origin at 440–370 Mya during the Silurian or Devonian. This discrepancy would require that winged insects evaded fossilization for at least the first ~50 Myr of their history. Here, we examine the plausibility of such a gap in the fossil record, and possible explanations for it, based on comparisons with the fossil records of other arthropod groups, the distribution of first occurrence dates of pterygote families, phylogenetically informed simulations of the fossilization of Palaeozoic insects, and re-analysis of data presented by Misof and colleagues using updated fossil calibrations under a variety of prior probability settings. We do not find support for the mechanisms previously suggested to account for such an extended gap in the pterygote fossil record, including sampling bias, preservation bias, and body size. We suggest that inference of an early origin of Pterygota long prior to their first appearance in the fossil record is probably an analytical artefact of taxon sampling and choice of fossil calibration points, possibly compounded by heterogeneity in rates of sequence evolution or speciation, including radiations or ‘bursts’ during their early history.

The uncertainity in stormwater quality modelling for temperate and tropical catchments

More research has been done on the uncertainty of stormwater quantity as compared to quality modelling. Although the washoff model has been in use for decades, there is a general lack of studies on the uncertainty associated with washoff modelling and how this uncertainty varies between catchments in different climate zones due to differences in rainfall conditions. This study investigated model structure uncertainty, in addition to gaining knowledge on model parameter uncertainty in water quality modelling of urban catchments located in two climate zones. Such a study not only adds to the body of knowledge but highlights the importance of distinguishing the difference in washoff behaviour, and their results, between catchments in different climate zones. Following an exhaustive search and assessment of data quality, the rainfall, discharge and water quality data sourced from 11 residential catchments, which include 72 events from temperate catchments and 59 from tropical catchments were analysed. Comparison of uncertainty between buildup-washoff model (BW) and washoff model (WO) results for particulate (Total Suspended Solids (TSS)), mixed (Total Phosphorus (TP), Total Nitrogen (TN)) and dissolved (ortho-Phosphate (OP), Ammonium (NH4-N)) phase water quality parameters were carried by using the Generalized Likelihood Uncertainty Estimation (GLUE) method. Higher Average Relative Interval Length (ARIL) values obtained for tropical catchments showed that uncertainty in model prediction is higher than temperate catchments. This result is confirmed for both the BW and WO modelling approaches. The threshold criteria for obtaining behavioural set chosen as likelihood measure of Tr = 0.7 (based on the Nash-Sutcliffe coefficient) and η ≥ 70 % (proportion of observed data falling within the 90 % uncertainty band of the simulated data distribution over the total number of observations) are mostly satisfied for the particulate phase and least by the dissolved phase water quality parameters, providing further evidence that the structure of the models are not universally applicable, and that model results should be understood in the light of this difference. Comparisons between the BW and WO approaches showed similar levels of uncertainty, irrespective of climate. Although this is consistent with other studies reported in the literature, the antecedent dry period (ADP), a key parameter in the buildup process, was not significantly (p =.07) different between the temperate and tropical datasets. This is likely an artifact of sampling, especially for temperate catchments, where field data is collected during the rainy season, when ADP is limited and similar with catchment in the tropics.

Gamma and electron beam irradiation use in the control of a radiation-resistant fungus (Epicoccum nigrum) for preservation of contaminated organic artefacts of traditional heritage

When artefacts of traditional cultural heritage are waterlogged due to floods and other disasters, they can be quickly protected from contamination by harmful fungi using irradiation treatments. In this study, we showed that Epicoccum nigrum has the potential to contaminate traditional heritage artefacts made of Korean paper and silk under waterlogged conditions. E. nigrum showed 60% lower mycelial growth in medium containing freshwater than in medium containing natural seawater. Control of this contamination was attempted using gamma and electron beam irradiation disinfection technology. The mycelial growth of E. nigrum irradiated with a 5 kGy dose of gamma rays was completely controlled at 15 kGy dose. Next, we evaluated the effect of irradiation on contaminated sample artefacts. E. nigrum growing on waterlogged sample artefacts of cultural heritage was completely controlled at 15 kGy in freshwater-immersed artefacts of Korean paper and silk and seawater-immersed silk. However, in Korean paper immersed in seawater, 15 kGy gamma-irradiation treatment did not control growth of the fungus. Our results suggest that at least 15 kGy of gamma radiation should be used to for treatment, to preserve waterlogged traditional cultural heritage artefacts of Korea, after flooding or other disasters involving water.

Laparoscopic twist technique has the best overall artifact profile when comparing three laparoscopic hepatic cup biopsy techniques for dogs.

To determine the difference in histologic artifacts and morphologic diagnosis among 3 laparoscopic cup biopsy forceps techniques and wedge hepatic samples. Cadavers of 20 client-owned dogs following euthanasia for unrelated reasons between January 3 and July 29, 2021. Four biopsy techniques were performed from the margin of 3 liver lobes/dog. Laparoscopic techniques used 5-mm cup biopsy forceps to obtain biopsy samples by pulling the forceps forcefully caudally to free a sample (the PULL technique), rotating the forceps 360° in 1 direction until freed (the TWST technique), or pulling the forceps through a 5-mm cannula to remove the sample (the CAN technique); wedge biopsy samples served as the control (CON). Data collected included sample weight, histologic features, diagnosis, and artifact characterization. Gwet AC1 or intraclass correlation coefficients (ICCs) were calculated to detect agreement among techniques. Sample weights for CON and TWST were significantly larger (P < .001 and P = .035, respectively) than for PULL and CAN. There was excellent agreement among all techniques for most diagnostic features (Gwet AC1, 0.93 to 1). The TWST technique resulted in the best overall artifact profile for laparoscopic techniques, with 90% of samples (54/60) having crisp edges and 65% of samples (39/60) having no or mild tearing. The agreement was moderate to good (ICC, 0.73 for edges and 0.76 for tearing) among all cup biopsy forceps techniques. The TWST technique resulted in the largest sample and had the fewest artifacts, supporting its continued use during laparoscopic procedures.

Comprehensive characterization of particulate intermediate-volatility and semi-volatile organic compounds (I/SVOCs) from heavy-duty diesel vehicles using two-dimensional gas chromatography time-of-flight mass spectrometry

Abstract. Tailpipe emissions from three heavy-duty diesel vehicles (HDDVs), complying with varying emission standards and installed with diverse aftertreatment technologies, are collected at a certified chassis dynamometer laboratory. The HDDV-emitted intermediate-volatility and semi-volatile organic compound (I/SVOC) emission and the gas–particle partitioning of the I/SVOCs are investigated. Over 4000 compounds are identified and grouped into 21 categories. The dominant compound groups of particulate I/SVOCs are alkanes and phenolic compounds. For HDDVs without aftertreatment devices, i.e., diesel oxidation catalysts (DOCs) and diesel particulate filters (DPFs), the emitted I/SVOCs partition dramatically into the gas phase (accounting for ∼ 93 % of the total I/SVOC mass), with a few exceptions: hopane, four-ring polycyclic aromatic hydrocarbons (PAH4rings), and five-ring polycyclic aromatic hydrocarbons (PAH5rings). For HDDVs with DPFs and DOCs, the particulate fractions are reduced to a negligible level (i.e., less than 2 %). Nevertheless, 50 % of the total two-ring PAH mass is detected in the particle phase, which is much higher than the high-molecular-weight PAHs, arising from the positive sampling artifact of quartz filter absorbing organic vapors. The positive sampling artifact of quartz filter absorbing organic vapors is clearly observed, and uncertainties are discussed and quantified. Particulate I/SVOCs at low-speed, middle-speed, and high-speed phases are collected and analyzed separately. The emission factor (EF) distribution of the speciated organic aerosol (OA) on a two-dimensional volatility basis set (2D-VBS) space reveals that the fractions of OA with oxygen to carbon (O : C) ratios &gt; 0.3 (0.4, 0.5) are 18.2 % (11.5 %, 9.5 %), 23 % (15.4 %, 13.6 %), and 29.1 % (20.6 %, 19.1 %) at the low-speed, middle-speed, and high-speed stages. These results help to resolve complex organic mixtures and trace the evolution of OA.

The development of diffusive equilibrium, high-resolution passive samplers to measure perfluoroalkyl substances (PFAS) in groundwater.

Per- and poly-fluoroalkyl substances (PFAS) are a group of anthropogenic, highly recalcitrant organic compounds consisting of thousands of individual species that are of increasing importance as groundwater contaminants. In-situ measurements of PFAS would be useful to better understand vertical profiles and mobility, contamination in partially saturated media, and to reduce sampling artifacts associated with groundwater collection and analysis. Diffusive equilibrium, high-resolution passive samplers (HRPPs) can be directly driven (>10m) in sediments or groundwater. The samplers equilibrate with porewater through diffusion across the sampler membrane, providing high spatial resolution (sample every 20cm) porewater concentrations of dissolved species. The objective of this study was to develop an HRPP to measure PFAS in contaminated groundwater and saturated media. To achieve this objective, a screening study was conducted to demonstrate quantitative measurement of selected PFAS as well as the kinetics of uptake into a sampler using both nylon and stainless steel membranes. Utilizing the results of the screening study, a prototype sampler was demonstrated in a laboratory flow box. Over a deployment period of 28 days, concentrations of several perfluoroalkyl carboxylic acids (PFCAs), a perfluoroalkyl sulfonate (PFSA), and a precursor PFAS reached equilibrium with porewater (sampler concentration >90 percent of porewater concentration). Application of these samplers could provide improved understanding of the behavior of PFAS in saturated or partially saturated groundwater systems and allow better assessment of fate and transport in the subsurface. Reliable subsurface site characterization will yield robust site assessments, conceptual models, and improve remediation designs as well as increase confidence in post remedial assessments at PFAS-impacted locations.

Scale space detector for analyzing spatiotemporal ventricular contractility and nuclear morphogenesis in zebrafish

SummaryIn vivo quantitative assessment of structural and functional biomarkers is essential for characterizing the pathophysiology of congenital disorders. In this regard, fixed tissue analysis has offered revolutionary insights into the underlying cellular architecture. However, histological analysis faces major drawbacks with respect to lack of spatiotemporal sampling and tissue artifacts during sample preparation. This study demonstrates the potential of light sheet fluorescence microscopy (LSFM) as a non-invasive, 4D (3days + time) optical sectioning tool for revealing cardiac mechano-transduction in zebrafish. Furthermore, we have described the utility of a scale and size-invariant feature detector, for analyzing individual morphology of fused cardiomyocyte nuclei and characterizing zebrafish ventricular contractility.

Assessment of perivascular space filtering methods using a three-dimensional computational model

Growing interest surrounds the assessment of perivascular spaces (PVS) on magnetic resonance imaging (MRI) and their validation as a clinical biomarker of adverse brain health. Nonetheless, the limits of validity of current state-of-the-art segmentation methods are still unclear. Here, we propose an open-source three-dimensional computational framework comprising 3D digital reference objects and evaluate the performance of three PVS filtering methods under various spatiotemporal imaging considerations (including sampling, motion artefacts, and Rician noise). Specifically, we study the performance of the Frangi, Jerman and RORPO filters in enhancing PVS-like structures to facilitate segmentation. Our findings were three-fold. First, as long as voxels are isotropic, RORPO outperforms the other two filters, regardless of imaging quality. Unlike the Frangi and Jerman filters, RORPO's performance does not deteriorate as PVS volume increases. Second, the performance of all “vesselness” filters is heavily influenced by imaging quality, with sampling and motion artefacts being the most damaging for these types of analyses. Third, none of the filters can distinguish PVS from other hyperintense structures (e.g. white matter hyperintensities, stroke lesions, or lacunes) effectively, the area under precision-recall curve dropped substantially (Frangi: from 94.21 [IQR 91.60, 96.16] to 43.76 [IQR 25.19, 63.38]; Jerman: from 94.51 [IQR 91.90, 95.37] to 58.00 [IQR 35.68, 64.87]; RORPO: from 98.72 [IQR 95.37, 98.96] to 71.87 [IQR 57.21, 76.63] without and with other hyperintense structures, respectively). The use of our computational model enables comparing segmentation methods and identifying their advantages and disadvantages, thereby providing means for testing and optimising pipelines for ongoing and future studies.

How innovations in methodology offer new prospects for volume electron microscopy

Detailed knowledge of biological structure has been key in understanding biology at several levels of organisation, from organs to cells and proteins. Volume electron microscopy (volume EM) provides high resolution 3D structural information about tissues on the nanometre scale. However, the throughput rate of conventional electron microscopes has limited the volume size and number of samples that can be imaged. Recent improvements in methodology are currently driving a revolution in volume EM, making possible the structural imaging of whole organs and small organisms. In turn, these recent developments in image acquisition have created or stressed bottlenecks in other parts of the pipeline, like sample preparation, image analysis and data management. While the progress in image analysis is stunning due to the advent of automatic segmentation and server‐based annotation tools, several challenges remain. Here we discuss recent trends in volume EM, emerging methods for increasing throughput and implications for sample preparation, image analysis and data management.

Evaluation of organic aerosol filter sampling artefacts and implications to gravimetric PM2.5 mass at a COALESCE network site - Bhopal, India

Filter based PM2.5 samples are typically used to measure its chemical constituents. Such measurements are made in dense sampling networks to assess regulatory compliance and for source apportionment. Thus, quantifying sampling artefacts is crucial. In this study, 24-h integrated PM2.5 samples collected over Bhopal, India a COALESCE (CarbOnaceous AerosoL Emissions, Source apportionment and ClimatE impacts) site during 2019 and 2020, were used to estimate particulate organic carbon (OC) artefacts. Total OC and its thermal fractions (OC1, OC2, OC3, and OC4) measured on 349 bare quartz (Q) and QbQ filters each, were used to determine OC positive artefacts on quartz filters. 50 QbT (Quartz behind Teflon) filters in conjunction with the simultaneous QbQ samples (a subset of the total QbQ) were used to estimate OC volatilization from Teflon filters. On average, adsorbed gaseous OC contributed 17% and 11% to the measured total OC during 2019 and 2020, respectively. Further, the volatilization loss of organics from Teflon filter (used to quantify PM2.5 mass) ranged between 7% and 9%, and 5% and 6% of the PM2.5 mass during 2019 and 2020, respectively. The results of this study provide the first systematic long-term evaluation of thermal carbon fraction-wise sampling artefacts, estimates of organic volatilization losses from Teflon filters and their implications to PM2.5 mass closure, over a regionally representative location in India.

Longitudinal Assessment of Contamination Patterns on Workstations on Wheels (WOWs)

Background Representations of contamination on portable medical equipment inform strategies to prevent transmission and healthcare-associated infections (HAIs). While periodic disinfection of equipment is required in most hospitals, the complexity of demands on staff may limit their ability to adequately disinfect multiple surfaces and touch points on a piece of equipment. The purpose of this study was to determine if current disinfection practices were effectively managing bioburden on WOWs on our hospital wards. Methods Samples were collected using press plate for four weeks from 10 WOWs on three medical-surgical units in our hospital. The sampling scheme limited the ability of staff to anticipate sampling and ensured multiple surfaces on each WOW were consistently sampled. Aerobic bacterial colony counts were calculated after 24 hours of incubation. Bayesian multilevel models estimated the mean bioburden and compared bioburden from different locations on WOWs. Results Out of the 452 samples, the estimated mean colony count (95% credible interval) was 29.2 (16.1-51.1). The incident risk-ratio comparing the handle to the other surfaces on the WOWs were 0.43 (0.32-0.55) for the keyboard, 0.29 (0.22-0.38) for the tray, and 0.22 (0.16-0.29) for the mouse. Conclusions The handle used for moving the WOW around, had greater bioburden compared to the keyboard, tray, or the mouse. Difference in the bioburden may reflect number of touches and lack of hand hygiene when moving equipment. The lower bioburden on keyboards and mouses may conversely reflect greater attention to hand hygiene or may be an artifact of sampling (less contact with press plates). The correlation of bioburden to HAI transmission was not studied here so the significance of bioburden alone in the absence of HAI monitoring is uncertain. Representations of contamination on portable medical equipment inform strategies to prevent transmission and healthcare-associated infections (HAIs). While periodic disinfection of equipment is required in most hospitals, the complexity of demands on staff may limit their ability to adequately disinfect multiple surfaces and touch points on a piece of equipment. The purpose of this study was to determine if current disinfection practices were effectively managing bioburden on WOWs on our hospital wards. Samples were collected using press plate for four weeks from 10 WOWs on three medical-surgical units in our hospital. The sampling scheme limited the ability of staff to anticipate sampling and ensured multiple surfaces on each WOW were consistently sampled. Aerobic bacterial colony counts were calculated after 24 hours of incubation. Bayesian multilevel models estimated the mean bioburden and compared bioburden from different locations on WOWs. Out of the 452 samples, the estimated mean colony count (95% credible interval) was 29.2 (16.1-51.1). The incident risk-ratio comparing the handle to the other surfaces on the WOWs were 0.43 (0.32-0.55) for the keyboard, 0.29 (0.22-0.38) for the tray, and 0.22 (0.16-0.29) for the mouse. The handle used for moving the WOW around, had greater bioburden compared to the keyboard, tray, or the mouse. Difference in the bioburden may reflect number of touches and lack of hand hygiene when moving equipment. The lower bioburden on keyboards and mouses may conversely reflect greater attention to hand hygiene or may be an artifact of sampling (less contact with press plates). The correlation of bioburden to HAI transmission was not studied here so the significance of bioburden alone in the absence of HAI monitoring is uncertain.

Subsurface microbial communities as a tool for characterizing regional-scale groundwater flow

Subsurface microbial community distribution patterns are influenced by biogeochemical and groundwater fluxes and may inform hydraulic connections along groundwater-flow paths. This study examined the regional-scale microbial community of the Death Valley Regional Flow System and evaluated whether subsurface communities can be used to identify groundwater-flow paths between recharge and discharge areas. Samples were collected from 36 sites in three groundwater basins: Pahute Mesa–Oasis Valley (PMOV), Ash Meadows (AM), and Alkali Flat–Furnace Creek Ranch (AFFCR). Microbial diversity within and between communities varied by location, and communities were separated into two overall groups that affiliated with the AM and PMOV/AFFCR basins. Network analysis revealed patterns between clusters of common microbes that represented groundwaters with similar geochemical conditions and largely corroborated hydraulic connections between recharge and discharge areas. Null model analyses identified deterministic and stochastic ecological processes contributing to microbial community assemblages. Most communities were more different than expected and governed by dispersal limitation, geochemical differences, or undominating processes. However, certain communities from sites located within or near the Nevada National Security Site were more similar than expected and dominated by homogeneous dispersal or selection. Overall, the (dis)similarities between the microbial communities of DVRFS recharge and discharge areas supported previously documented hydraulic connections between: (1) Spring Mountains and Ash Meadows; (2) Frenchman and Yucca Flat and Amargosa Desert; and (3) Amargosa Desert and Death Valley. However, only a portion of the flow path between Pahute Mesa and Oasis Valley could be supported by microbial community analyses, likely due to well-associated artifacts in samples from the two Oasis Valley sites. This study demonstrates the utility of combining microbial data with hydrologic, geologic, and water-chemistry information to comprehensively characterize groundwater systems, highlighting both strengths and limitations of this approach.

Abstract 2185: MicroSEC: Sequence error filter for formalin-fixed and paraffin-embedded samples

Abstract Background: DNAs extracted from formalin-fixed and paraffin-embedded (FFPE) tissues are fragmented and contain a significant amount of single-stranded DNA (ssDNA). We found that most of the false-positive mutations in clinical sequencing with FFPE samples are ssDNA-derived artifacts. During the end-repair step of library preparation, chimeric reads are generated by mis-annealing of ssDNA molecules comprising homologous sequences with a mismatched base. The mismatched base is called as a false-positive mutation, and the position of the base is biased towards the effective ends in the individual reads because either side of the chimeric read should be soft-clipped by the mapping pipeline. Based on this theory, we developed a post hoc filter, MicroSEC, to predict such artifacts in FFPE samples. Methods: Fifty-three fresh frozen (FF) and 190 FFPE normal breast tissue samples and 23 FF and 33 FFPE breast cancer samples were obtained from 26 patients, and subjected to capture-based panel sequencing (Todai OncoPanel, TOP). We also obtained TOP data from 54 FFPE various tumor samples. MicroSEC was developed to predict artifacts with BAM files based on the positional bias of mutations within each read. The predictions were validated with amplicon-based sequencing of 97 mutations. We also developed a model which predicts artifacts only from the mutated base information without corresponding BAM files, by using 5,034 MicroSEC predictions as the supervised data with the LightGBM technique. A total of 742,030 mutations from the AACR Project GENIE were examined with the model. Results: Among the normal breast tissues, we identified 0.3 and 11.7 somatic mutations per sample in FF and FFPE specimens, and 0 (0%) and 10.1 (86.0%) were filtered out by MicroSEC, respectively. Two unique mutations with variant allele frequency of &amp;gt;50% in FFPE samples were eliminated. From the breast cancer specimens, we identified 4.0 and 10.7 mutations per sample in FF and FFPE, and 0 (0%) and 3.2 (30.2%) were filtered out, respectively. In the clinical sequencing data of 54 FFPE tumor samples, we identified 21.6 mutations per sample and 3.6 (16.6%) including five unique pathogenic mutations were filtered out. The validation study showed that the sensitivity and specificity for artifacts of MicroSEC were 97% (95% confidence interval (CI): 82%-100%) and 96% (95% CI: 88%-99%), respectively. Further, the prediction model reproduced the MicroSEC predictions with an area under the ROC curve of 0.995. The model detected 2,512 mutations (0.34%) as artifacts from Project GENIE data. There was a difference in the artifact detection rate between institutions, and 815 (1.45%) of the 56,100 mutations reported by UCSF were predicted as artifacts. Conclusions: MicroSEC removes only FFPE artifacts without eliminating true mutations found in FF samples. Our pipeline will increase the reliability of the clinical sequencing and advance cancer research using FFPE samples. Citation Format: Masachika Ikegami, Shinji Kohsaka, Takeshi Hirose, Toshihide Ueno, Satoshi Inoue, Naoki Kanomata, Hideko Yamauchi, Taisuke Mori, Shigeki Sekine, Yoshihiro Inamoto, Yasushi Yatabe, Hiroshi Kobayashi, Sakae Tanaka, Toru Akiyama, Tomotake Okuma, Hiroyuki Mano. MicroSEC: Sequence error filter for formalin-fixed and paraffin-embedded samples [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2185.

Comparison and evaluation of enumeration methods for measurement of fungal spore emission

Estimation of fungal spore emission from various sources, which is a necessary input for the estimation of their effects on health and climate using mathematical models, relies on realistic quantification of fungal spores. Different quantification techniques have been used in various studies of fungal spore emission. For the purposes of model development, validation and application, the various methods must be correlated. In this study, we compare the commonly used methods - fluorescence microscopy, haemocytometer and fluorescence spectroscopy, in in relation to fungal spore emission estimation using Penicillium chrysogenum as a test fungal species. In addition, we also compare an offline and online technique - Biosampler® and Optical particle sizer (OPS) respectively. It was found that haemocytometer shows up to 50% higher average spore count than fluorescence microscopy in the range of 104–107 spores/ml. The effect of sampling volume, artifacts and image magnification on sample representativeness was also studied using fluorescence microscopy as the basis. Statistical analysis of these effects shows that at spore concentrations lower than 104/mL, the uncertainty is significant. The comparison between emission measurement using OPS and Biosampler/fluorescence microscopy techniques were reasonably similar. The quantitative comparison also provides a basis for other users to choose methods for their research objectives using appropriate quantitative correction and correlation factors.

Reducing pre-analytical sample QC failure rates for cancer molecular genetic assays with SLIMamp technology.

e15034 Background: Identification of somatic variants in cancer by high-throughput sequencing has become common clinical practice largely because many of these variants may be predictive biomarkers for targeted therapies. However, there can be high sample QC failure rates for some assays (sometimes up to 40%), preventing the return of results that may affect patient treatment decisions. Pillar Biosciences has incorporated their patented SLIMamp technology into commercially available cancer NGS testing kits with the claim that these kits can successfully interrogate challenging formalin-fixed paraffin-embedded tissue (FFPET) samples with low tumor purity, poor DNA quality, and/or low input DNA, resulting in a high sample QC pass rate. The aim of this study was to substantiate that claim using Pillar’s amplicon-based oncoReveal Solid Tumor Panel. Methods: We acquired 48 tumor samples that had failed one or more pre-analytical QC sample parameters for whole exome sequencing (WES) from ATGC’s ISO15189 accredited diagnostic genomics laboratory. XING Genomic Services performed an exploratory data analysis using our pre-analytical QC assays to characterise the samples and then sequenced the samples in our ISO15189 accredited laboratory using the validated oncoReveal Solid Tumor Panel. Results: We were able to achieve high sequencing coverage (&gt;3000X) for all 48 samples and explored the determinants of sample “success”. We were able to generate clinical reports for 45 samples (94%), of which 38 (79%) contained clinically actionable or significant variants that would not have otherwise been identified. Ten samples had a higher number of total variant calls with over-representation of C&gt;T transitions representing stochastically-amplified, formalin-induced artefacts in samples with very low input template DNA. Of these, 7 cases had reportable variants and 3 were deemed unreportable. We demonstrated that DNA integrity is the major determinant of success even in samples with low input DNA or low tumor purity and were able to further refine pre-analytical and post-analytical QC metrics to better identify samples with poor quality DNA that can be sequenced reliably. Conclusions: In this study, we showed that the Pillar Biosciences oncoReveal Solid Tumor Panel, which uses SLIMam technology, was able to generate reliable, interpretable results for 94% of samples that failed pre-analytical QC for WES, substantiating Pillar’s claim.

Evaluating bone biopsy quality by technique in an animal model.

Powered bone biopsy technique is popular due to its ease of use. However, there is conflicting evidence regarding the diagnostic quality of the samples. The purpose of this study is to evaluate the diagnostic adequacy of different bone biopsy devices and techniques as it relates to the frequency of sample artifacts. Bone biopsy was performed on same-day processed lamb femora using the following techniques: manual, pulsed powered and full powered. Ten samples were collected using each method by a single musculoskeletal-trained radiologist and were reviewed by 3 blinded pathologists. Samples were compared across multiple categories: length, bone dust, thermal/crush artifact, cellular morphology, fragmentation, and diagnostic acceptability. Bayesian Multilevel Nonlinear Regression models were performed assessing the association between the techniques across the categories. Statistical analysis revealed that the manual technique outperformed any powered technique across all categories: decreased thermal/crush artifact (P = 0.014), decreased bone dust (p<0.001), better cellular morphology (P = 0.005), less fragmentation (P < 0.0001) and better diagnostic acceptability (P < 0.0001). Manually obtained bone biopsy samples generally produce a more diagnostic sample as compared to powered techniques in an animal model. Given these results, manual bone biopsy methods should be encouraged after consideration for lesion composition, difficulty of access and the patient's overall condition.

Vacuum-Formed 3D Printed Electronics: Fabrication of Thin, Rigid and Free-Form Interactive Surfaces

Vacuum-forming is a common manufacturing technique for constructing thin plastic shell products by pressing heated plastic sheets onto a mold using atmospheric pressure. Vacuum-forming is ubiquitous in packaging and casing products in the industry, spanning fast moving consumer goods to connected devices. Integrating advanced functionality, which may include sensing, computation and communication, within thin structures is desirable for various next-generation interactive devices. Hybrid additive manufacturing techniques like thermoforming are becoming popular for prototyping freeform surfaces owing to their design flexibility, speed and cost-effectiveness. This paper presents a new hybrid method for constructing thin, rigid and free-form interconnected surfaces via fused deposition modelling (FDM) 3D printing and vacuum-forming that builds on recent advances in thermoforming circuits. 3D printing the sheet material allows for the embedding of conductive traces within thin layers of the substrate, which can be vacuum-formed but remain conductive and insulated. This is an unexplored fabrication technique within the context of designing and manufacturing connected things. In addition to explaining the method, this paper characterizes the behavior of vacuum-formed 3D printed sheets, analyses the electrical performance of printed traces after vacuum-forming, and showcases a range of sample artefacts constructed using the technique. In addition, the paper describes a new design interface for designing conformal interconnects that allows designers to draw conductive patterns in 3D and export pre-distorted sheet models ready to be printed.

The contribution of canopy samples to assessments of forestry effects on native bees

AbstractForest management is often practiced to enhance conditions for wildlife, including native bees. Evaluations of the effects of forest management on bees have shown that abundance and diversity are higher in newly created early‐successional conditions. To date, studies have restricted sampling to the forest understory; however, recent research finds that bee abundance is as high or higher in forest canopies than in understories, suggesting that previous observations of substantially greater bee abundance and diversity in recently managed areas could be an artifact of incomplete sampling of the vertical gradient within forests. To examine the potential implications of sampling biases associated with the failure of previous studies to include canopy samples in comparisons of managed and unmanaged forests, we sampled bees within a recently harvested forest as well as the understory and canopy of adjacent unmanaged forest. Bee abundance and diversity were an order of magnitude higher in managed areas compared to the unmanaged forest, even when understory and canopy samples were combined. These results suggest that not sampling the canopy is inconsequential with respect to the broadly reported conclusion that managed areas support more abundant and diverse bee communities than surrounding forest cover.

Checklist, diversity and biogeography of ciliates (Ciliophora) from Chile

There is a significant gap in research and knowledge on the diversity and distribution of Chilean ciliates. To tackle these issues, we used cultures and protargol preparations to describe the ciliates present in poorly explored areas. At these sites, we identified 45 ciliate morphospecies, 35 of which represent unprecedent records to Chile. Then, we brought together our records with literature data to construct a species checklist. This checklist summarises 132 years of data and describes the identity, habitat and distribution of 207 species, including 15 species potentially endemic to Chile. This checklist is far from complete: a diversity estimate suggests that at least two-thirds of the ciliate species occurring in Chile have yet to be described. The checklist is dominated by freshwater taxa because ciliates from marine, brackish and terrestrial environments have rarely been investigated in Chile. Finally, after controlling for sampling artefacts, we found that ciliates exhibit a bell-shaped latitudinal diversity gradient in Chile. This peculiar biogeographical pattern is common in Chile. Plants, animals and testate amoebae also exhibit a bell-shaped latitudinal diversity gradient in Chile. This finding suggests that the historical contingencies that drove the biogeography of the Chilean biota also shaped ciliate biogeography.

A Novel Step-by-Step Assessment Methodology for the Correction of Sampling Artifact and Chlorine Loss Reaction on HNO3 in Ambient Air

A Novel Step-by-Step Assessment Methodology for the Correction of Sampling Artifact and Chlorine Loss Reaction on HNO3 in Ambient Air