Linear systems characterization of the topographical spatial resolution of optical instruments.

Lateral resolving power is a key performance attribute of Fizeau interferometers, confocal microscopes, interference microscopes, and other instruments measuring surface form and texture. Within a well-defined scope of applicability, limited by surface slope, texture, and continuity, a linear response model provides a starting point for characterizing spatial resolution under ideal conditions. Presently, the instrument transfer function (ITF) is a standardized way to quantify linear response to surface height variations as a function of spatial frequency. In this paper, we build on the ITF idea and introduce terms, mathematical definitions, and appropriate physical units for applying a linear systems model to surface topography measurement. These new terms include topographical equivalents of the point-, line-, and edge-spread functions, as well as a complex-valued transfer function that extends the ITF concept to systems with spatial-frequency-dependent topography distortions. As an example, we consider the experimental determination of lateral resolving power of a coherence scanning interference microscope using a step-height surface feature to measure the ITF directly. The experiment illustrates the proposed mathematical definitions and provides a direct comparison to theoretical calculations performed using a scalar diffraction model.

Non-invasive biomonitoring of polar bear feces can be used to estimate concentrations of metals of concern in traditional food.

The Arctic faces increasing exposure to environmental chemicals such as metals, posing health risks to humans and wildlife. Biomonitoring of polar bears (Ursus maritimus) can be used to quantify chemicals in the environment and in traditional foods consumed by the Inuit. However, typically, these samples are collected through invasive or terminal methods. The biomonitoring of feces could be a useful alternative to the current metal monitoring method within the Arctic. Here, we aim to 1) quantify the relationship between concentrations of metals in the feces and tissues (muscle, liver, and fat) of polar bears using predictive modeling, 2) develop an easy-to-use conversion tool for use in community-based monitoring programs to non-invasively estimate contaminant concentrations in polar bears tissues and 3) demonstrate the application of these models by examining potential exposure risk for humans from consumption of polar bear muscle. Fecal, muscle, liver, and fat samples were harvested from 49 polar bears through a community-based monitoring program. The samples were analyzed for 32 metals. Exploratory analysis indicated that mean metal concentrations generally did not vary by age or sex, and many of the metals measured in feces were positively correlated with the internal tissue concentration. We developed predictive linear regression models between internal (muscle, liver, fat) and external (feces) metal concentrations and further explored the mercury and methylmercury relationships for utility risk screening. Using the cross-validated regression coefficients, we developed a conversion tool that contributes to the One Health approach by understanding the interrelated health of humans, wildlife, and the environment in the Arctic. The findings support using feces as a biomonitoring tool for assessing contaminants in polar bears. Further research is needed to validate the developed models for other regions in the Arctic and assess the impact of environmental weathering on fecal metal concentrations.

The geographic distribution, and the biotic and abiotic predictors of select zoonotic pathogen detections in Canadian polar bears.

Increasing Arctic temperatures are facilitating the northward expansion of more southerly hosts, vectors, and pathogens, exposing naïve populations to pathogens not typical at northern latitudes. To understand such rapidly changing host-pathogen dynamics, we need sensitive and robust surveillance tools. Here, we use a novel multiplexed magnetic-capture and droplet digital PCR (ddPCR) tool to assess a sentinel Arctic species, the polar bear (Ursus maritimus; n = 68), for the presence of five zoonotic pathogens (Erysipelothrix rhusiopathiae, Francisella tularensis, Mycobacterium tuberculosis complex, Toxoplasma gondii and Trichinella spp.), and observe associations between pathogen presence and biotic and abiotic predictors. We made two novel detections: the first detection of a Mycobacterium tuberculosis complex member in Arctic wildlife and the first of E. rhusiopathiae in a polar bear. We found a prevalence of 37% for E. rhusiopathiae, 16% for F. tularensis, 29% for Mycobacterium tuberculosis complex, 18% for T. gondii, and 75% for Trichinella spp. We also identify associations with bear age (Trichinella spp.), harvest season (F. tularensis and MTBC), and human settlements (E. rhusiopathiae, F. tularensis, MTBC, and Trichinella spp.). We demonstrate that monitoring a sentinel species, the polar bear, could be a powerful tool in disease surveillance and highlight the need to better characterize pathogen distributions and diversity in the Arctic.

Population genomics of the muskox' resilience in the near absence of genetic variation.

Genomic studies of species threatened by extinction are providing crucial information about evolutionary mechanisms and genetic consequences of population declines and bottlenecks. However, to understand how species avoid the extinction vortex, insights can be drawn by studying species that thrive despite past declines. Here, we studied the population genomics of the muskox (Ovibos moschatus), an Ice Age relict that was at the brink of extinction for thousands of years at the end of the Pleistocene yet appears to be thriving today. We analysed 108 whole genomes, including present-day individuals representing the current native range of both muskox subspecies, the white-faced and the barren-ground muskox (O. moschatus wardi and O. moschatus moschatus) and a ~21,000-year-old ancient individual from Siberia. We found that the muskox' demographic history was profoundly shaped by past climate changes and post-glacial re-colonizations. In particular, the white-faced muskox has the lowest genome-wide heterozygosity recorded in an ungulate. Yet, there is no evidence of inbreeding depression in native muskox populations. We hypothesize that this can be explained by the effect of long-term gradual population declines that allowed for purging of strongly deleterious mutations. This study provides insights into how species with a history of population bottlenecks, small population sizes and low genetic diversity survive against all odds.

A new multiplexed magnetic capture-Droplet digital PCR tool for monitoring wildlife population health and pathogen surveillance.

Anthropogenic stressors are exacerbating the emergence and spread of pathogens worldwide. In regions like the Arctic, where ecosystems are particularly susceptible, marked changes are predicted in regional diversity, intensity, and patterns of infectious diseases. To understand such rapidly changing host-pathogen dynamics and mitigate the impacts of novel pathogens, we need sensitive disease surveillance tools. We developed and validated a novel multiplexed, magnetic capture, and ddPCR tool for the surveillance of multiple pathogens in polar bears, a sentinel species that is considered susceptible to climate change and other stressors with a pan-Arctic distribution. Through sequence-specific magnetic capture, we concentrated five target template sequences from three zoonotic bacteria (Erysipelothrix rhusiopathiae, Francisella tularensis, and Mycobacterium tuberculosis complex) and two parasitic (Toxoplasma gondii and Trichinella spp.) pathogens from large quantities (<100 g) of host tissue. We then designed and validated two multiplexed probe-based ddPCR assays for the amplification and detection of the low-concentration target DNA. Validations used 48 polar bear tissues (muscle and liver). We detected 14, 1, 3, 4, and 22 tissue positives for E. rhusiopathiae, F. tularensis, M. tuberculosis complex, T. gondii, and Trichinella spp., respectively. These multiplexed assays offer a rapid, specific tool for quantifying and monitoring the changing geographical and host distributions of pathogens relevant to human and animal health.

Outcomes of hyperbolic tapering of antidepressants.

Background: In patients attempting to discontinue their antidepressant medication, there have been no prospective studies on patterns of withdrawal as a function of the rate of antidepressant reduction during the tapering trajectory, and moderators thereof. Objective: To investigate withdrawal as a function of gradual dose reduction. Design: Prospective cohort study. Methods: The sampling frame consisted of 3956 individuals in the Netherlands who received an antidepressant tapering strip between 19 May 2019 and 22 March 2022 in routine clinical practice. Of these, 608 patients, majorly with previous unsuccessful attempts to stop, provided daily ratings of withdrawal in the context of reducing their antidepressant medications (mostly venlafaxine or paroxetine), using hyperbolic tapering strips offering daily tiny reductions in dose. Results: Withdrawal in daily-step hyperbolic tapering trajectories was limited, and inverse to the rate of taper. Female sex, younger age, presence of one or more risk factors and faster rate of reduction over shorter tapering trajectories were associated with more withdrawal and differential course over time. Thus, sex and age differences were less marked early in the course of the trajectory, whereas differences associated with risk factors and shorter trajectories tended to peak early in the trajectory. There was evidence that tapering in weekly larger steps (mean per-week dose reduction: 33.4% of previous dose), in comparison with daily tiny steps (mean per-day dose reduction: 4.5% of previous dose or 25.3% per week), was associated with more withdrawal in trajectories of 1, 2 or 3 months, particularly for paroxetine and the group of other (non-paroxetine, non-venlafaxine) antidepressants. Conclusion: Antidepressant hyperbolic tapering is associated with limited, rate-dependent withdrawal that is inverse to the rate of taper. The demonstration of multiple demographic, risk and complex temporal moderators in time series of withdrawal data indicates that antidepressant tapering in clinical practice requires a personalised process of shared decision making over the entire course of the tapering period.

Long-Term Mild Heat Causes Post-Mitotic Pollen Abortion Through a Local Effect on Flowers.

Crop reproductive success is significantly challenged by heatwaves, which are increasing in frequency and severity globally. Heat-induced male sterility is mainly due to aborted pollen development, but it is not clear whether this is through direct or systemic effects. Here, long-term mild heat (LTMH) treatment, mimicking a heatwave, was applied locally to tomato flowers or whole plants and followed up by cytological, transcriptomic, and biochemical analyses. By analyzing pollen viability, LTMH was shown to act directly on the flowers and not via effects on other plant tissue. The meiosis to early microspore stage of pollen development was the most sensitive to LTMH and 3 days of exposure around this period was sufficient to significantly reduce pollen viability at the flower anthesis stage. Extensive cytological analysis showed that abnormalities in pollen development could first be observed after pollen mitosis I, while no deviations in tapetum development were observed. Transcriptomic and biochemical analyses suggested that pollen development suffered from tapetal ER stress and that there was a limited role for oxidative stress. Our results provide the first evidence that heat acts directly on flowers to induce pollen sterility, and that the molecular-physiological responses of developing anthers to the LTMH are different from those to severe heat shock.

Genotyping-in-thousands by sequencing (GT-seq) of noninvasive faecal and degraded samples: A new panel to enable ongoing monitoring of Canadian polar bear populations.

Genetic monitoring using noninvasive samples provides a complement or alternative to traditional population monitoring methods. However, next-generation sequencing approaches to monitoring typically require high quality DNA and the use of noninvasive samples (e.g., scat) is often challenged by poor DNA quality and contamination by nontarget species. One promising solution is a highly multiplexed sequencing approach called genotyping-in-thousands by sequencing (GT-seq), which can enable cost-efficient genomics-based monitoring for populations based on noninvasively collected samples. Here, we develop and validate a GT-seq panel of 324 single nucleotide polymorphisms (SNPs) optimized for genotyping of polar bears based on DNA from noninvasively collected faecal samples. We demonstrate (1) successful GT-seq genotyping of DNA from a range of sample sources, including successful genotyping (>50% loci) of 62.9% of noninvasively collected faecal samples determined to contain polar bear DNA; and (2) that we can reliably differentiate individuals, ascertain sex, assess relatedness, and resolve population structure of Canadian polar bear subpopulations based on a GT-seq panel of 324 SNPs. Our GT-seq data reveal spatial-genetic patterns similar to previous polar bear studies but at lesser cost per sample and through use of noninvasively collected samples, indicating the potential of this approach for population monitoring. This GT-seq panel provides the foundation for a noninvasive toolkit for polar bear monitoring and can contribute to community-based programmes - a framework which may serve as a model for wildlife conservation and management for species worldwide.

Surface measuring coherence scanning interferometry beyond the specular reflection limit.

The capability of optical surface topography measurement methods for measurement of steep and tilted surfaces is investigated through modelling of a coherence scanning interferometer. Of particular interest is the effect on the interference signal and measured topography when tilting the object at angles larger than the numerical aperture slope limit (i.e. the specular reflection limit) of the instrument. Here we use theoretical modelling to predict the results across a range of tilt angles for a blazed diffraction grating. The theoretically predicted interference patterns and surface height measurements are then verified directly with experimental measurements. Results illustrate the capabilities, limitations and modelling methods for interferometers to measure beyond the specular reflection limit.

Correlation of Mercury Occurrence with Age, Elemental Composition, and Life History in Sea-Run Food Fish from the Canadian Arctic Archipelago's Lower Northwest Passage.

As mercury emissions continue and climate-mediated permafrost thaw increases the burden of this contaminant in northern waters, Inuit from a Northwest passage community in the Canadian Arctic Archipelago pressed for an assessment of their subsistence catches. Sea-run salmonids (n = 537) comprising Arctic char (Salvelinus alpinus), lake trout (S. namaycush), lake whitefish (Coregonus clupeaformis), and cisco (C. autumnalis, C. sardinella) were analyzed for muscle mercury. Methylmercury is a neurotoxin and bioaccumulated with fish age, but other factors including selenium and other elements, diet and trophic level as assessed by stable isotopes of nitrogen (δ15N) and carbon (δ13C), as well as growth rate, condition, and geographic origin, also contributed depending on the species, even though all the fish shared a similar anadromous or sea-run life history. Although mean mercury concentrations for most of the species were ~0.09 µg·g-1 wet weight (ww), below the levels described in several jurisdictions for subsistence fisheries (0.2 µg·g-1 ww), 70% of lake trout were above this guideline (0.35 µg·g-1 ww), and 19% exceeded the 2.5-fold higher levels for commercial sale. We thus urge the development of consumption advisories for lake trout for the protection of pregnant women and young children and that additionally, periodic community-based monitoring be initiated.

Successful use of tapering strips for hyperbolic reduction of antidepressant dose: a cohort study.

BACKGROUND: Tapering strips facilitate antidepressant discontinuation, allowing for personalised titration of discontinuation to the intensity of withdrawal. A tapering strip consists of antidepressant or other medication, packaged in a 28-day roll of daily pouches, each with the same or slightly lower dose than the one before. Previous studies demonstrated 70% real-world effectiveness of tapering strips. Here, we present a third, questionnaire-based retrospective cohort study in a large sample. METHODS: Patients whose doctor had prescribed tapering strips between October 2015 and December 2018 were sent a questionnaire for participation after completion of tapering between December 2015 and January 2020. Of 1240 individuals who returned a questionnaire (response rate: 59%), 987 (80%) used an antidepressant, of whom 824 (83%) had wished to discontinue their antidepressant. RESULTS: The sample was demographically representative of antidepressant users in the Netherlands. Less than 40% of participants had heard of tapering strips through their clinicians - Internet was the most frequent source. Of the 824 individuals, 341 (41%) had used strips for tapering venlafaxine, 206 (25%) for paroxetine and 277 (34%) for other antidepressants. Median duration of antidepressant use was 5-10 years, and most (71%) had tried to come off without tapering strips at least once. Most patients (72%) were able to discontinue their antidepressant, using a median of two strips to taper over a median period of 56 days. Females and individuals with (1) more severe experience of withdrawal during the use of tapering strips, (2) more years of use of antidepressant medication and (3) more previous attempts at discontinuation were less likely to be able to discontinue their antidepressant medication with tapering strips. CONCLUSION: The results of this study validate, for the third time, the observation that tapering strips can address the problem of antidepressant withdrawal symptoms in individuals attempting to discontinue antidepressants.

Free-Living, Psychrotrophic Bacteria of the Genus Psychrobacter Are Descendants of Pathobionts.

Host-adapted microorganisms are generally assumed to have evolved from free-living, environmental microorganisms, as examples of the reverse process are rare. In the phylum Gammaproteobacteria, family Moraxellaceae, the genus Psychrobacter includes strains from a broad ecological distribution including animal bodies as well as sea ice and other nonhost environments. To elucidate the relationship between these ecological niches and Psychrobacter's evolutionary history, we performed tandem genomic analyses with phenotyping of 85 Psychrobacter accessions. Phylogenomic analysis of the family Moraxellaceae reveals that basal members of the Psychrobacter clade are Moraxella spp., a group of often-pathogenic organisms. Psychrobacter exhibited two broad growth patterns in our phenotypic screen: one group that we called the "flexible ecotype" (FE) had the ability to grow between 4 and 37°C, and the other, which we called the "restricted ecotype" (RE), could grow between 4 and 25°C. The FE group includes phylogenetically basal strains, and FE strains exhibit increased transposon copy numbers, smaller genomes, and a higher likelihood to be bile salt resistant. The RE group contains only phylogenetically derived strains and has increased proportions of lipid metabolism and biofilm formation genes, functions that are adaptive to cold stress. In a 16S rRNA gene survey of polar bear fecal samples, we detect both FE and RE strains, but in in vivo colonizations of gnotobiotic mice, only FE strains persist. Our results indicate the ability to grow at 37°C, seemingly necessary for mammalian gut colonization, is an ancestral trait for Psychrobacter, which likely evolved from a pathobiont.IMPORTANCE Host-associated microbes are generally assumed to have evolved from free-living ones. The evolutionary transition of microbes in the opposite direction, from host associated toward free living, has been predicted based on phylogenetic data but not studied in depth. Here, we provide evidence that the genus Psychrobacter, particularly well known for inhabiting low-temperature, high-salt environments such as sea ice, permafrost soils, and frozen foodstuffs, has evolved from a mammalian-associated ancestor. We show that some Psychrobacter strains retain seemingly ancestral genomic and phenotypic traits that correspond with host association while others have diverged to psychrotrophic or psychrophilic lifestyles.

Production Time and User Satisfaction of 3-Dimensional Printed Orthoses For Chronic Hand Conditions Compared With Conventional Orthoses: A Prospective Case Series.

OBJECTIVE: Hand orthoses are often prescribed for persons with chronic hand and wrist impairments. This study assessed the feasibility, in terms of production time and user satisfaction, of 3-dimensional printed hand orthoses compared with conventional hand orthoses for this population. METHODS: In this prospective case series, both a conventional hand orthosis and a 3-dimensional printed hand orthosis were manufactured for 10 participants. Production time (in minutes) of each orthosis was recorded. Each orthosis was worn for one week, after which participants completed a self-designed questionnaire on satisfaction, scored on a 5-point Likert scale. Functionality and orthosis preference were also assessed. RESULTS: The mean (standard deviation (SD)) production time for the 3-dimensional printed orthoses, of 112 (11.0) min, was significantly shorter compared with 239 (29.2) min for the conventional orthoses (95% confidence interval (95% CI) 71-182 min, p = 0.001). Satisfaction scores were similar for both orthoses, except for comfort item "fitting method", which was rated significantly higher for scanning compared with casting (median [IQR] score: 5 [0.0]; 4 [2.0], p = 0.034). Functionality and orthosis preference were rated similar for both orthoses. CONCLUSION: As the production time was halved, user satisfaction similar, and scanning experienced as slightly more comfortable than casting, 3-dimensional printed hand orthoses seem feasible and potentially beneficial for use in people with chronic hand and wrist impairments.

Characterization of 29 polymorphic microsatellite markers developed by genomic screening of Sumatran rhinoceros (Dicerorhinus sumatrensis).

OBJECTIVE: The Sumatran rhinoceros is critically endangered, with fewer than 100 individuals surviving across its current range. Accurate census estimates of the remaining populations are essential for development and implementation of conservation plans. In order to enable molecular censusing, we here develop microsatellite markers with amplicon sizes of short length, appropriate for non-invasive fecal sampling. RESULTS: Due to limited sample quantity and potential lack of genome-wide diversity, Illumina sequence reads were generated from two Sumatran rhinoceros samples. Genomic screening identified reads with short tandem repeats and loci that were polymorphic within the dataset. Twenty-nine novel polymorphic microsatellite markers were characterized (A = 2.4; HO = 0.30). These were sufficient to distinguish among individuals (PID < 0.0001), and to distinguish among siblings (PID(sib) < 0.0001). Among rhinos in Indonesia, almost all markers were established as polymorphic and effective for genotyping DNA from fecal samples. Notably, the markers amplified and displayed microsatellite polymorphisms using DNA extracted from 11 fecal samples collected non-invasively from wild Sumatran rhinoceros. These microsatellite markers provide an important resource for a census and genetic studies of wild Sumatran rhinos.

Outcome of antidepressant drug discontinuation with taperingstrips after 1-5 years.

BACKGROUND: Stopping antidepressants is often difficult due to withdrawal. Taperingstrips were developed to facilitate antidepressant discontinuation according to the recently described Horowitz-Taylor method, allowing for personalised titration of discontinuation to the intensity of withdrawal. A taperingstrip consists of antidepressant or other medication, packaged in a 28-day roll of small daily pouches, each with the same or slightly lower dose than the one before it. We previously reported that the short-term success rate of antidepressant taperingstrips was 71%. Here, we examine longer-term outcome after 1-5 years. METHODS: Patients whose doctor had ordered taperingstrips between January 2015 and December 2019 were sent a questionnaire for participation in anonymised research in January 2020. Of 1012, 483 participated, of whom 408 (85%) had attempted antidepressant tapering. RESULTS: Of the 408 patients included, 192 (47%) had used strips for tapering venlafaxine, 142 (35%) for paroxetine and 74 (18%) for other antidepressants. Median length of antidepressant use was 4 years, and most (61%) had tried to come off without taperingstrips at least once. After 1-5 years, 270 patients (66%) remained off antidepressants after tapering their antidepressant, 6 (2%) had successfully reduced their medication, 87 (21%) had restarted due to (self-reported) relapse, 35 had restarted for another indication (9%), and 10 (3%) reported another outcome. People with more severe experience of withdrawal prior to tapering, and people who had been on antidepressants for a shorter period of time, were more likely to remain off medication after 1-5 years. CONCLUSION: The previously reported 71% short-term success rate of taperingstrips in the most severely affected group, was matched by a 68% rate after 1-5 years. The evidence-based approach of personal tapering to counter withdrawal, as used for drugs causing withdrawal, for example, benzodiazepines, may represent a simple solution for an important antidepressant-related public health problem, without extra costs.

Fourier optics modeling of interference microscopes.

We propose a practical theoretical model of an interference microscope that includes the imaging properties of optical systems with partially coherent illumination. We show that the effects on measured topography of a spatially extended, monochromatic light source at low numerical apertures can be approximated in a simplified model that assumes spatially coherent light and a linear, locally shift-invariant transfer function that accounts for optical aberrations and the attenuation of diffracted plane wave amplitudes with increasing spatial frequencies. Simulation of instrument response using this model agrees with methods using numerical pupil-plane integration and with an experimental measurement of surface topography.

How user knowledge of psychotropic drug withdrawal resulted in the development of person-specific tapering medication.

Coming off psychotropic drugs can cause physical as well as mental withdrawal, resulting in failed withdrawal attempts and unnecessary long-term drug use. The first reports about withdrawal appeared in the 1950s, but although patients have been complaining about psychotropic withdrawal problems for decades, the first tentative acknowledgement by psychiatry only came in 1997 with the introduction of the 'antidepressant-discontinuation syndrome'. It was not until 2019 that the UK Royal College of Psychiatrists, for the first time, acknowledged that withdrawal can be severe and persistent. Given the lack of a systematic professional response, over the years, patients who were experiencing withdrawal started to work out practical ways to safely come off medications themselves. This resulted in an experience-based knowledge base about withdrawal which ultimately, in The Netherlands, gave rise to the development of person-specific tapering medication (so-called tapering strips). Tapering medication enables doctors, for the first time, to flexibly prescribe and adapt the medication required for responsible and person-specific tapering, based on shared decision making and in full agreement with recommendations in existing guidelines. Looking back, it is obvious that the simple practical solution of tapering strips could have been introduced much earlier, and that the traditional academic strategy of comparisons from randomised trials is not the logical first step to help individual patients. While randomised controlled trials (RCTs) are the gold standard for evaluating interventions, they are unable to accommodate the heterogeneity of individual responses. Thus, a more individualised approach, building on RCT knowledge, is required. We propose a roadmap for a more productive way forward, in which patients and academic psychiatry work together to improve the recognition and person-specific management of psychotropic drug withdrawal.

Seasonal habitat drives intestinal microbiome composition in anadromous Arctic char (Salvelinus alpinus).

Intestinal microbial communities from 362 anadromous Arctic char (Salvelinus alpinus) from the high Arctic Kitikmeot region, Nunavut, Canada, were characterized using high-throughput 16S rRNA gene sequencing. The resulting bacterial communities were compared across four seasonal habitats that correspond to different stages of annual migration. Arctic char intestinal communities differed by sampling site, salinity and stages of freshwater residence. Although microbiota from fish sampled in brackish water were broadly consistent with taxa seen in other anadromous salmonids, they were enriched with putative psychrophiles, including the nonluminous gut symbiont Photobacterium iliopiscarium that was detected in >90% of intestinal samples from these waters. Microbiota from freshwater-associated fish were less consistent with results reported for other salmonids, and highly variable, possibly reflecting winter fasting behaviour of these char. We identified microbiota links to age for those fish sampled during the autumn upriver migration, but little impact of the intestinal content and water microbiota on the intestinal community. The strongest driver of intestinal community composition was seasonal habitat, and this finding combined with identification of psychrophiles suggested that water temperature and migratory behaviour are key to understanding the relationship between Arctic char and their symbionts.

A real-time PCR assay to accurately quantify polar bear DNA in fecal extracts.

DNA extracted from fecal samples contains DNA from the focal species, food, bacteria and pathogens. Most DNA quantification methods measure total DNA and cannot differentiate among sources. Despite the desirability of noninvasive fecal sampling for studying wildlife populations, low amounts of focal species DNA make it difficult to use for next-generation sequencing (NGS), where accurate DNA quantification is critical for normalization. Two factors are required prior to using fecal samples in NGS libraries: (1) an accurate quantification method for the amount of target DNA and (2) a determination of the relative amount of target DNA needed for successful single nucleotide polymorphism genotyping assays. Here, we address these needs by developing primers to amplify a 101 bp region of the nuclear F2 gene and a quantitative PCR (qPCR) assay that allows the accurate quantification of the amount of polar bear (Ursus maritimus) DNA in fecal extracts. We test the assay on pure polar bear DNA extracted from muscle tissue and find a high correlation between fluorometric and qPCR quantifications. The qPCR assay was also successfully used to quantify the amount of DNA derived from polar bears in fecal extractions. Orthologs of the F2 gene have been identified across vertebrates; thus, similar qPCR assays could be developed for other species to enable noninvasive studies.