A GPS assisted translocation experiment to study the homing behavior of red deer.

Many animals return to their home areas (i.e., 'homing') after translocation to sites further away. Such translocations have traditionally been used in behavioral ecology to understand the orientation and migration behavior of animals. The movement itself can then be followed by marking and recapturing animals or by tracking, for example, using GPS systems. Most detailed studies investigating this behavior have been conducted in smaller vertebrates (e.g., birds, amphibians, and mice), whereas information on larger mammals, such as red deer, is sparse. We conducted GPS-assisted translocation experiments with red deer at two sites in the Czech Republic. Individuals were translocated over a distance of approximately 11 km and their home journey was tracked. Circular statistics were used to test for significant homeward orientation at distances of 100, 500, 1000, and 5000 m from the release site. In addition, we applied Lavielle trajectory segmentation to identify the different phases of homing behavior. Thirty-one out of 35 translocations resulted in successful homing, with a median time of 4.75 days (range 1.23-100 days). Animals were significantly oriented towards home immediately after release and again when they came closer to home; however, they did not show a significant orientation at the distances in between. We were able to identify three homing phases, an initial 'exploratory phase', followed by a 'homing phase' which sometimes was again followed by an 'arrival phase'. The 'homing phase' was characterized by the straightest paths and fastest movements. However, the variation between translocation events was considerable. We showed good homing abilities of red deer after translocation. Our results demonstrate the feasibility of conducting experiments with environmental manipulations (e.g., to impede the use of sensory cues) close to the release site. The homing behavior of red deer is comparable to that of other species, and might represent general homing behavior patterns in animals. Follow-up studies should further dissect and investigate the drivers of the individual variations observed and try to identify the sensory cues used during homing.

'Phototaxis' in the absence of light? Locomotory patterns in unionid mussels.

Locomotion in benthic invertebrates can strongly affect habitat selection and ecosystem nutrient cycling. In the case of freshwater mussels, the drivers of locomotion are largely unresolved. Our aim was to assess the influence of light presence and intensity on the locomotory behaviour of freshwater mussels in controlled laboratory experiments. The species investigated in our study were Anodonta anatina and Unio pictorum, two widely distributed mussels in European lentic and lotic inland waters. At low algal concentrations, known to be associated with more frequent locomotory activities, we found that both species moved primarily in the absence of light (72.7% of all movements across experiments). However, the movements of both species were directed towards the light source, resembling a net-positive 'phototactic' response but in the absence of light. The distance to the light source, which was negatively correlated to light intensity, had a positive effect on the distance covered in locomotory activities by A. anatina but not by U. pictorum. Intraspecific variation in shell size had no impact on movement distance, indicating that the energetic costs of movement were not a limiting factor. We suggest that the observed movement towards brighter locations helps to enhance food quantity and quality, whilst movement in darkness mitigates predation risks.

Evaluating the power of a recent method for comparing two circular distributions: an alternative to the Watson U2 test.

Some data are collected on circular (rather than linear) scales. Often researchers are interested in comparing two samples of such circular data to test the hypothesis that they came from the same underlying population. Recently, we compared 18 statistical approaches to testing such a hypothesis, and recommended two as particularly effective. A very recent publication introduced a novel statistical approach that was claimed to outperform the methods that we had indicated were highest performing. However, the evidence base for this claim was limited. Here we perform simulation studies to offer a more detailed comparison of the new "Angular Randomisation Test" (ART) with existing tests. We expand previous evaluations in two ways: exploring small and medium sized samples, and exploring a range of different shapes for the underlying distribution(s). We find that the ART controls type I error rates at the nominal level. The ART had greater power than established methods in detecting a difference in underlying distribution caused by a shift around the circle. Its performance advantage in this case was strongest when samples where small and unbalanced in size. When the difference between underlying unimodal distributions was in shape rather than central tendency, then the ART was at least as good (and sometimes considerably more powerful) than the established methods, except when distributions samples were small and uneven in size, and the smaller sample came from a more concentrated underlying distribution. In such cases its power could be markedly inferior to established alternatives. The ART was also inferior to alternatives in dealing with axially distributed data. We conclude that under widely-encountered circumstances the ART test can be recommended for its simplicity of implementation, but researchers should be aware of situations where it cannot be recommended.

Telemetry and Accelerometer Tracking of Green Toads in an Urban Habitat: Methodological Notes and Preliminary Findings.

Advancements in tracking technologies provide an increasingly important tool in animal monitoring and conservation that can describe animal spatial behavior in native habitats and uncover migratory routes that otherwise may be difficult or impossible to map. In addition, high-resolution accelerometer sensors provide powerful insights into animal activity patterns and can help to identify specific behaviors from accelerometer profiles alone. Previously, such accelerometers were restricted to larger animals due to size and mass constraints. However, recent advances make it possible to use such devices on smaller animals such as the European green toad (Bufotes viridis), the focus of our current study. We deploy custom made tracking devices, that consist of very-high-frequency transmitters and tri-axial accelerometers, to track toads in their native urban environment in Vienna (Austria). A total of nine toads were tracked, ranging from three to nine tracking days per individual during the post-breeding season period. We demonstrate that our devices could reliably monitor toad movement and activity during the observation period. Hence, we confirmed the predominantly nocturnal activity patterns and recorded low overall movement at this urban site. Accelerometer data revealed that toads exhibited brief but intense activity bursts between 10 pm and midnight, resting periods during the night and intermittent activity during the day. Positional tracking alone would have missed the major activity events as they rarely resulted in large positional displacements. This underscores the importance of and value in integrating multiple tracking sensors for studies of movement ecology. Our approach could be adapted for other amphibians or other animals with mass constraints and may become standard monitoring equipment in the near future.

Temporal and spatial variations in local sex ratios in a suburban population of the European green toad Bufotes viridis.

BACKGROUND: Sex ratios of animal populations are important factors of population demographics. In pond-breeding amphibians, the operational sex ratio (OSR) among the breeding population is usually male-biased. Also, in European green toads (Bufotes viridis), males usually outnumber females at breeding sites, while the sex ratio of the total adult population (ASR) is assumed to be balanced. It has been suggested that sex-specific breeding behavior causes male-predominance at the breeding sites. We used a dataset of 5 years of street patrols to test this hypothesis. For this we analyzed local sex ratios of green toads in terrestrial habitats and at two artificial breeding ponds. We expected temporal and/or spatial changes of local sex ratios which would indicate sex dependent differences in breeding behavior. RESULTS: Overall observed ASR among 2111 green toads, counted in the course of street patrols from 2016 to 2020, was slightly male-biased (ASR = 0.56, annual ASRs = 0.49-0.63). Based on the data of 1631 toads (920 males, 711 females) captured within a radius of 300 m around nine main breeding sites, temporal and spatial variations in local ASRs were evaluated. Resulting values were compared to the calculated OSR at two artificial breeding ponds in 2021 (645 adult: 553 males, 92 females). Estimates predict more equally distributed females and males prior to the main breeding season. During breeding season, males predominated at both breeding sites (B1: 0.83, B2: 0.89), whereas females are estimated to outnumber males in terrestrial habitats. Proportions of females highly significantly increased with advancing time of the year and increasing distance to the breeding sites. While males tended to accumulate in proximity to water bodies, females dispersed soon after breeding to more distant areas. CONCLUSIONS: Observed sex ratios in the studied green toad population changed with time and sampling site, deviating from the population-wide sex ratio. Expanding sampling effort in amphibian conservation assessments in time and space, i.e., outside the main breeding season and away from the breeding sites, would be important to encompass such variations.

Land-use preferences of the European green toad (Bufotes viridis) in the city of Vienna (Austria): the importance of open land in urban environments.

Urban areas are increasing worldwide, which poses threats to animal wildlife. However, in certain cases cities can provide refuges for endangered animals. The European green toad (Bufotes viridis) is one of such examples, which is known from cities throughout their distribution. In contrast, considerable areas of their former (primary) habitats have been degraded. The primary habitats of this species include steppes and wild river floodplains, both characterized by dynamic changes and the presence of open areas. We used available green toad observation data (2007-2020) to model the effects of land-use types on occurrence probability in the city of Vienna. Forest and densely populated areas were highly significantly negatively associated with green toad presence, while transformation/construction site areas showed a strong positive effect. Such occurrence pattern might be characteristic for early succession species, which depend on stochastic environmental disturbances (e.g., droughts and floods) in their primary habitats. We argue that urban landscape planning should appreciate the potential ecological value of open land in cities which is either in a transition phase or a permanent 'wasteland'. Ecological managing of such landscape could vastly increase urban biodiversity.

Unexpected air pollutants with potential human health hazards: Nitrification inhibitors, biocides, and persistent organic substances.

To better understand the influence of land use and meteorological parameters on air pollutants, we deployed passive air samplers in 15 regions with different land use in eastern Austria. The samplers consisted of polyurethane PUF and polyester PEF filter matrices, which were analyzed for 566 substances by gas-chromatography/mass-spectrometry. In a previous article, we highlighted a widespread contamination of ambient air with pesticides that depends on the surrounding land use and meteorological parameters. Here we report that, in addition to agricultural pesticides, eight other substances were frequently detected in ambient air: Nitrapyrin, a nitrification inhibitor used to increase nitrogen use efficiency of fertilizers and banned in Austria since 1993; biocides against insects (DEET and transfluthrin) used mainly outside agriculture; piperonyl butoxide (PBO), a synergist mixed into pesticide formulations; and four industrially used polychlorinated biphenyls (PCBs), long banned worldwide. Concentrations of the detected substances were positively related to air temperature, but only slightly related to agricultural land use in the sampler's vicinity. The city center showed the highest concentrations of biocides, PCBs and PBO, but also medium concentrations of nitrapyrin. Four sites had no air contamination with these substances; including two national parks dominated by grassland or forest, but also two sites with mixed land use. The potential human toxicity of the detected substances based on globally harmonized hazard classifications was high: seven substances had specific organ toxicity, six were cancerogenic, and two were acutely toxic; however, several substances had incomplete information of hazard profiles. Moreover, all substances were acutely and chronically toxic to aquatic life. We recommend that substances of different origins be included in the air pollution monitoring portfolio to comprehensively assess the potential hazards to humans and the environment.

Effects of cavity orientation on nesting success inferred from long-term monitoring of the endangered red-cockaded woodpecker.

Animals that create structures often display non-random patterns in the direction of their constructions. This tendency of oriented construction is widely presumed to be an adaptive trait of the constructor's extended phenotype, but there is little empirical support for this hypothesis. Particularly, for cavity nesting-birds there is a lack of studies examining this issue. In this study of a primary cavity excavator, the endangered red-cockaded woodpecker (Dryobates borealis), we show that cavity entrances exhibited a strong westward bias in all 11 of the populations examined throughout the geographic range of the species in the southeastern United States. This species requires cavities in living pine trees for roosting and nesting that often take many years to complete, resulting in many incomplete excavations on the landscape. We used population monitoring data to show that orientation was stronger among completed cavities than incomplete cavities. There was a significant correlation between latitude and average cavity direction among populations, turning northward with increasing latitude, suggesting adaptation to local conditions. Long-term monitoring data showed that cavity orientation and breeding group size are correlated with egg hatching rates, fledging rates, and the total number of fledglings produced per nest. Our results provide empirical evidence from extensive long-term data that directional orientation in animal constructions is an important feature of the extended animal phenotype and have immediate implications for animal ecology and the conservation of endangered species.

Pesticides in ambient air, influenced by surrounding land use and weather, pose a potential threat to biodiversity and humans.

Little is known about (i) how numbers and concentrations of airborne pesticide residues are influenced by land use, interactions with meteorological parameters, or by substance-specific chemo-physical properties, and (ii) what potential toxicological hazards this could pose to non-target organisms including humans. We installed passive air samplers (polyurethane PUF and polyester PEF filter matrices) in 15 regions with different land uses in eastern Austria for up to 8 months. Samples were analyzed for 566 substances by gas-chromatography/mass-spectrometry. We analyzed relationships between frequency and concentrations of pesticides, land use, meteorological parameters, substance properties, and season. We found totally 67 pesticide active ingredients (24 herbicides, 30 fungicides, 13 insecticides) with 10-53 pesticides per site. Herbicides metolachlor, pendimethalin, prosulfocarb, terbuthylazine, and the fungicide HCB were found in all PUF samplers, and glyphosate in all PEF samplers; chlorpyrifos-ethyl was the most abundant insecticide found in 93% of the samplers. Highest concentrations showed the herbicide prosulfocarb (725 ± 1218 ng sample-1), the fungicide folpet (412 ± 465 ng sample-1), and the insecticide chlorpyrifos-ethyl (110 ± 98 ng sample-1). Pesticide numbers and concentrations increased with increasing proportions of arable land in the surroundings. However, pesticides were also found in two National Parks (10 and 33 pesticides) or a city center (17 pesticides). Pesticide numbers and concentrations changed between seasons and correlated with land use, temperature, radiation, and wind, but were unaffected by substance volatility. Potential ecotoxicological exposure of mammals, birds, earthworms, fish, and honeybees increased with increasing pesticide numbers and concentrations. Human toxicity potential of detected pesticides was high, with averaged 54% being acutely toxic, 39% reproduction toxic, 24% cancerogenic, and 10% endocrine disrupting. This widespread pesticide air pollution indicates that current environmental risk assessments, field application techniques, protective measures, and regulations are inadequate to protect the environment and humans from potentially harmful exposure.

Orientation and emigration of larval and juvenile amphibians: selected topics and hypotheses.

Most amphibians have a complex life cycle with an aquatic larval and an adult (semi-) terrestrial stage. However, studies concerning spatial behaviour and orientation mainly focus on either the aquatic larvae or the adult animals on land. Consequently, behavioural changes that happen during metamorphosis and the consequences for emigration and population distribution are less understood. This paper aims to summarize the knowledge concerning specific topics of early amphibian life history stages and proposes several testable hypotheses within the following fields of research: larval and juvenile orientation, influences of environmental and genetic factors on juvenile emigration, their habitat choice later in life as well as population biology. I argue that studying larval and juvenile amphibian spatial behaviour is an understudied field of research, however, could considerably improve our understanding of amphibian ecology.

The multivariate analysis of variance as a powerful approach for circular data.

BACKGROUND: A broad range of scientific studies involve taking measurements on a circular, rather than linear, scale (often variables related to times or orientations). For linear measures there is a well-established statistical toolkit based on linear modelling to explore the associations between this focal variable and potentially several explanatory factors and covariates. In contrast, statistical testing of circular data is much simpler, often involving either testing whether variation in the focal measurements departs from circular uniformity, or whether a single explanatory factor with two levels is supported. METHODS: We use simulations and example data sets to investigate the usefulness of a MANOVA approach for circular data in comparison to commonly used statistical tests. RESULTS: Here we demonstrate that a MANOVA approach based on the sines and cosines of the circular data is as powerful as the most-commonly used tests when testing deviation from a uniform distribution, while additionally offering extension to multi-factorial modelling that these conventional circular statistical tests do not. CONCLUSIONS: The herein presented MANOVA approach offers a substantial broadening of the scientific questions that can be addressed statistically using circular data.

Why is it so difficult to study magnetic compass orientation in murine rodents?

A magnetic compass sense has been demonstrated in all major classes of vertebrates, as well as in many invertebrates. In mammals, controlled laboratory studies of mice have provided evidence for a robust magnetic compass that is comparable to, or exceeds, the performance of that in other animals. Nevertheless, the vast majority of laboratory studies of spatial behavior and cognition in murine rodents have failed to produce evidence of sensitivity to magnetic cues. Given the central role that a magnetic compass sense plays in the spatial ecology and cognition of non-mammalian vertebrates, and the potential utility that a global/universal reference frame derived from the magnetic field would have in mammals, the question of why responses to magnetic cues have been so difficult to demonstrate reliably is of considerable importance. In this paper, we review evidence that the magnetic compass of murine rodents shares a number of properties with light-dependent compasses in a wide variety of other animals generally believed to be mediated by a radical pair mechanism (RPM) or related quantum process. Consistent with the RPM, we summarize both published and previously unpublished findings suggesting that the murine rodent compass is sensitive to low-level radio frequency (RF) fields. Finally, we argue that the presence of anthropogenic RF fields in laboratory settings, may be an important source of variability in responses of murine rodents to magnetic cues.

A Unified Approach to Analysis of Body Condition in Green Toads.

Body condition is increasingly used to assess the status of populations and as a proxy for individual fitness. A common, quick and non-invasive approach is to estimate condition from the relation between body length and mass. Among the methods developed for this purpose, the Scaled Mass Index (SMI) appears best suited for comparisons among populations. We assembled data from 17 populations of European green toads (Bufotes viridis) with the aim of devising a standard formula applicable for monitoring this species. The mean value of the exponents describing length-mass allometry in these samples was 3.0047. Hence, we propose using 3 as a scaling coefficient for calculating the SMI in green toads. From the contrast of SMI values for both sexes within populations, estimated with either the population-specific or the standard coefficient, we conclude that applying the standard formula not only facilitates comparisons among populations but may also help to avoid misinterpretation of variation within populations.

Advice on comparing two independent samples of circular data in biology.

Many biological variables are recorded on a circular scale and therefore need different statistical treatment. A common question that is asked of such circular data involves comparison between two groups: Are the populations from which the two samples are drawn differently distributed around the circle? We compared 18 tests for such situations (by simulation) in terms of both abilities to control Type-I error rate near the nominal value, and statistical power. We found that only eight tests offered good control of Type-I error in all our simulated situations. Of these eight, we were able to identify the Watson's U2 test and a MANOVA approach, based on trigonometric functions of the data, as offering the best power in the overwhelming majority of our test circumstances. There was often little to choose between these tests in terms of power, and no situation where either of the remaining six tests offered substantially better power than either of these. Hence, we recommend the routine use of either Watson's U2 test or MANOVA approach when comparing two samples of circular data.

Long-term monitoring of common spadefoot toad activity in a European steppe using barn owl pellets.

BACKGROUND: One third of the worldwide amphibian species are threatened, therefore, efficient monitoring efforts are needed. Amphibians which adopt a hidden lifestyle, such as the common spadefoot toad, are often missed with standard surveying efforts. Spadefoot toads can be identified in regurgitated pellets of the barn owl, which provides an effective way to estimate toad activity. In our study we analyzed frequency of spadefoot toad remains from 2004 to 2016 in a steppe landscape in eastern Austria. METHODS: We used an automated model selection procedure together with a GLM analysis using a zero inflated error Poisson distribution, to analyze the presence of Pelobates fuscus in barn owl pellets. All analyses were done in the statistical software R, and the scripts to reproduce our results are available within this publication. Our approach may provide a template for other researchers to use for their own pellet data. CONCLUSIONS: Our analysis suggested that activity of the common spadefoot toad is mainly influenced by rainfalls, while time of the year and temperature had small but significant effects. Interestingly, our data confirmed the possibility of a second breeding period in summer, triggered by heavy rainfalls. There were no indications for a population decrease in the observed years and locations. Our study shows that barn owl pellets can be used effectivley to assess pelobatid activity in an area. This might constitute a useful monitoring tool for conservation management for amphibians.

The European green toad, Bufotes viridis, in Donaufeld (Vienna, Austria): status and size of the population.

The European green toad, Bufotes viridis (Laurenti, 1768), is a rare and protected species in Vienna. In spring and summer 2020, we conducted a survey to assess size and status of its population in Donaufeld, an agricultural area designated for real estate development. Recaptures of photographically registered toads allowed to estimate the population size with 137 individuals (confidence interval: 104-181). Comparatively large body size indicates the presence of a well-established population. Reproductive success was high in the study year. A mismatch mating of a male B. viridis with a female Bufo bufo was observed. Mitigation measures are needed to support this population facing imminent habitat deterioration.

A proteomic survey of microtubule-associated proteins in a R402H TUBA1A mutant mouse.

Microtubules play a critical role in multiple aspects of neurodevelopment, including the generation, migration and differentiation of neurons. A recurrent mutation (R402H) in the α-tubulin gene TUBA1A is known to cause lissencephaly with cerebellar and striatal phenotypes. Previous work has shown that this mutation does not perturb the chaperone-mediated folding of tubulin heterodimers, which are able to assemble and incorporate into the microtubule lattice. To explore the molecular mechanisms that cause the disease state we generated a new conditional mouse line that recapitulates the R402H variant. We show that heterozygous mutants present with laminar phenotypes in the cortex and hippocampus, as well as a reduction in striatal size and cerebellar abnormalities. We demonstrate that homozygous expression of the R402H allele causes neuronal death and exacerbates a cell intrinsic defect in cortical neuronal migration. Microtubule sedimentation assays coupled with quantitative mass spectrometry demonstrated that the binding and/or levels of multiple microtubule associated proteins (MAPs) are perturbed by the R402H mutation including VAPB, REEP1, EZRIN, PRNP and DYNC1l1/2. Consistent with these data we show that the R402H mutation impairs dynein-mediated transport which is associated with a decoupling of the nucleus to the microtubule organising center. Our data support a model whereby the R402H variant is able to fold and incorporate into microtubules, but acts as a gain of function by perturbing the binding of MAPs.

The biophysical, molecular, and anatomical landscape of pigeon CRY4: A candidate light-based quantal magnetosensor.

The biophysical and molecular mechanisms that enable animals to detect magnetic fields are unknown. It has been proposed that birds have a light-dependent magnetic compass that relies on the formation of radical pairs within cryptochrome molecules. Using spectroscopic methods, we show that pigeon cryptochrome clCRY4 is photoreduced efficiently and forms long-lived spin-correlated radical pairs via a tetrad of tryptophan residues. We report that clCRY4 is broadly and stably expressed within the retina but enriched at synapses in the outer plexiform layer in a repetitive manner. A proteomic survey for retinal-specific clCRY4 interactors identified molecules that are involved in receptor signaling, including glutamate receptor-interacting protein 2, which colocalizes with clCRY4. Our data support a model whereby clCRY4 acts as an ultraviolet-blue photoreceptor and/or a light-dependent magnetosensor by modulating glutamatergic synapses between horizontal cells and cones.

Grouped circular data in biology: advice for effectively implementing statistical procedures.

ABSTRACT: The most common statistical procedure with a sample of circular data is to test the null hypothesis that points are spread uniformly around the circle without a preferred direction. An array of tests for this has been developed. However, these tests were designed for continuously distributed data, whereas often (e.g. due to limited precision of measurement techniques) collected data is aggregated into a set of discrete values (e.g. rounded to the nearest degree). This disparity can cause an uncontrolled increase in type I error rate, an effect that is particularly problematic for tests that are based on the distribution of arc lengths between adjacent points (such as the Rao spacing test). Here, we demonstrate that an easy-to-apply modification can correct this problem, and we recommend this modification when using any test, other than the Rayleigh test, of circular uniformity on aggregated data. We provide R functions for this modification for several commonly used tests. In addition, we tested the power of a recently proposed test, the Gini test. However, we concluded that it lacks sufficient increase in power to replace any of the tests already in common use. In conclusion, using any of the standard circular tests (except the Rayleigh test) without modifications on rounded/aggregated data, especially with larger sample sizes, will increase the proportion of false-positive results-but we demonstrate that a simple and general modification avoids this problem. SIGNIFICANCE STATEMENT: Circular data are widespread across biological disciplines, e.g. in orientation studies or circadian rhythms. Often these data are rounded to the nearest 1-10 degrees. We have shown previously that this leads to an inflation of false-positive results when testing whether the data is significantly different from a random distribution using the Rao test. Here we present a modification that avoids this increase in false-positives for rounded data while retaining statistical power for a variety of tests. In sum, we provide comprehensive guidance on how best to test for departure from uniformity in non-continuous data.

Model selection versus traditional hypothesis testing in circular statistics: a simulation study.

Many studies in biology involve data measured on a circular scale. Such data require different statistical treatment from those measured on linear scales. The most common statistical exploration of circular data involves testing the null hypothesis that the data show no aggregation and are instead uniformly distributed over the whole circle. The most common means of performing this type of investigation is with a Rayleigh test. An alternative might be to compare the fit of the uniform distribution model to alternative models. Such model-fitting approaches have become a standard technique with linear data, and their greater application to circular data has been recently advocated. Here we present simulation data that demonstrate that such model-based inference can offer very similar performance to the best traditional tests, but only if adjustment is made in order to control type I error rate.