Cascading effects of earthworm invasion increase graminoid density and rodent grazing intensities.

Human-mediated dispersal of non-native earthworms can cause substantial changes to the functioning and composition of ecosystems previously earthworm-free. Some of these earthworm species have the potential to "geoengineer" soils and increase plant nitrogen (N) uptake. Yet the possible consequences of increased plant N concentrations on rodent grazing remains poorly understood. In this study, we present findings from a common garden experiment with two tundra communities, meadow (forb dominated) and heath (shrub dominated), half of them subjected to 4 years of earthworm presence (Lumbricus spp. and Aporrectodea spp.). Within four summers, our earthworm treatment changed plant community composition by increasing graminoid density by, on average, 94% in the heath vegetation and by 49% in the meadow. Rodent winter grazing was more intense on plants growing in soils with earthworms, an effect that coincided with higher N concentrations in plants, indicating a higher palatability. Even though earthworms reduced soil moisture, plant community productivity, as indicated by vegetation greenness (normalized difference vegetation index), was not negatively impacted. We conclude that earthworm-induced changes in plant composition and trophic interactions may fundamentally alter the functioning of tundra ecosystems.

The topological nature of tag jumping in environmental DNA metabarcoding studies.

Metabarcoding of environmental DNA constitutes a state-of-the-art tool for environmental studies. One fundamental principle implicit in most metabarcoding studies is that individual sample amplicons can still be identified after being pooled with others-based on their unique combinations of tags-during the so-called demultiplexing step that follows sequencing. Nevertheless, it has been recognized that tags can sometimes be changed (i.e., tag jumping), which ultimately leads to sample crosstalk. Here, using four DNA metabarcoding data sets derived from the analysis of soils and sediments, we show that tag jumping follows very specific and systematic patterns. Specifically, we find a strong correlation between the number of reads in blank samples and their topological position in the tag matrix (described by vertical and horizontal vectors). This observed spatial pattern of artefactual sequences could be explained by polymerase activity, which leads to the exchange of the 3' tag of single stranded tagged sequences through the formation of heteroduplexes with mixed barcodes. Importantly, tag jumping substantially distorted our data sets-despite our use of methods suggested to minimize this error. We developed a topological model to estimate the noise based on the counts in our blanks, which suggested that 40%-80% of the taxa in our soil and sedimentary samples were likely false positives introduced through tag jumping. We highlight that the amount of false positive detections caused by tag jumping strongly biased our community analyses.

Non-native species change the tune of tundra soils: Novel access to soundscapes of the Arctic earthworm invasion.

Over the last decade, an increasing number of studies have used soundscapes to address diverse ecological questions. Sound represents one of the few sources of information capable of providing in situ insights into processes occurring within opaque soil matrices. To date, the use of soundscapes for soil macrofauna monitoring has been experimentally tested only in controlled laboratory environments. Here we assess the validity of laboratory predictions and explore the use of soil soundscape proxies for monitoring soil macrofauna (i.e., earthworm) activities in an outdoor context. In a common garden experiment in northern Sweden, we constructed outdoor mesocosm plots (N = 36) containing two different Arctic vegetation types (meadow and heath) and introduced earthworms to half of these plots. Earthworms substantially altered the ambient soil soundscape under both vegetation types, as measured by both traditional soundscape indices and frequency band power levels, although their acoustic impacts were expressed differently in heath versus meadow soils. While these findings support the as-of-yet untapped promise of using belowground soundscape analyses to monitor soil ecosystem health, direct acoustic emissions from earthworm activities appear to be an unlikely proxy for tracking worm activities at daily timescales. Instead, earthworms indirectly altered the soil soundscape by 're-engineering' the soil matrix: an effect that was dependent on vegetation type. Our findings suggest that long-term (i.e., seasonal) earthworm activities in natural soil settings can likely be monitored indirectly via their impacts on soundscape measures and acoustic indices. Analyzing soil soundscapes may enable larger-scale monitoring of high-latitude soils and is directly applicable to the specific case of earthworm invasions within Arctic soils, which has recently been identified as a potential threat to the resilience of high-latitude ecosystems. Soil soundscapes could also offer a novel means to monitor soils and soil-plant-faunal interactions in situ across diverse pedogenic, agronomic, and ecological systems.

Norway spruce postglacial recolonization of Fennoscandia.

Contrasting theories exist regarding how Norway spruce (Picea abies) recolonized Fennoscandia after the last glaciation and both early Holocene establishments from western microrefugia and late Holocene colonization from the east have been postulated. Here, we show that Norway spruce was present in southern Fennoscandia as early as 14.7 ± 0.1 cal. kyr BP and that the millennia-old clonal spruce trees present today in central Sweden likely arrived with an early Holocene migration from the east. Our findings are based on ancient sedimentary DNA from multiple European sites (N = 15) combined with nuclear and mitochondrial DNA analysis of ancient clonal (N = 135) and contemporary spruce forest trees (N = 129) from central Sweden. Our other findings imply that Norway spruce was present shortly after deglaciation at the margins of the Scandinavian Ice Sheet, and support previously disputed finds of pollen in southern Sweden claiming spruce establishment during the Lateglacial.

Sodium hypochlorite as an oxidizing agent for removal of soil organic matter before microplastics analyses.

The omnipresence of microplastics (MPs) across Earth's surface has raised concerns about their environmental impact and created an urgent need for methods to identify them in complex soil and sedimentary matrices. However, detecting MPs in the O horizons of soils is difficult because plastic polymers share many physical and chemical properties with natural soil organic matter (SOM). In this study, we assessed whether sodium hypochlorite (NaOCl), a reagent that can oxidize SOM and simultaneously preserve mineral constituents, can be used for MP analysis and characterization in soil environments. In addition, we scrutinized how factors such as MP size, polymer type, extraction methods, and soil matrix affect the recovery of microplastic particles. We used both hydrophobic and density-dependent separation methods to assess the effects of our oxidation treatment on the recovery of MP. We observed that NaOCl effectively removed SOM without greatly altering the surface properties of resistant MP polymers (polypropylene, polylactic acid, low-density polyethylene, and polyethylene terephthalate), which were characterized using scanning electron microscopy and Fourier-transform infrared spectroscopy after SOM removal. The NaOCl treatment caused some chlorination and formation of additional C-OH bonds on polymer surfaces, which likely contributed to the reduced efficiency of the hydrophobic-based (oil) extraction. We conclude that NaOCl treatment can improve detection of MPs in SOM-rich soil and that recovery of MPs from soils is influenced by MP size, polymer type, extraction method, and soil type, which makes it challenging to develop a universal analytical method.

Exposure via biotransformation: Oxazepam reaches predicted pharmacological effect levels in European perch after exposure to temazepam.

It is generally expected that biotransformation and excretion of pharmaceuticals occurs similarly in fish and mammals, despite significant physiological differences. Here, we exposed European perch (Perca fluviatilis) to the benzodiazepine drug temazepam at a nominal concentration of 2 µg L-1 for 10 days. We collected samples of blood plasma, muscle, and brain in a time-dependent manner to assess its bioconcentration, biotransformation, and elimination over another 10 days of depuration in clean water. We observed rapid pharmacokinetics of temazepam during both the exposure and depuration periods. The steady state was reached within 24 h of exposure in most individuals, as was complete elimination of temazepam from tissues during depuration. Further, the biologically active metabolite oxazepam was produced via fish biotransformation, and accumulated significantly throughout the exposure period. In contrast to human patients, where a negligible amount of oxazepam is created by temazepam biotransformation, we observed a continuous increase of oxazepam concentrations in all fish tissues throughout exposure. Indeed, oxazepam accumulated more than its parent compound, did not reach a steady state during the exposure period, and was not completely eliminated even after 10 days of depuration, highlighting the importance of considering environmental hazards posed by pharmaceutical metabolites.

Impacts of Oxazepam on Perch (Perca fluviatilis) Behavior: Fish Familiarized to Lake Conditions Do Not Show Predicted Anti-anxiety Response.

A current theory in environmental science states that dissolved anxiolytics (oxazepam) from wastewater effluents can reduce anti-predator behavior in fish with potentially negative impacts on prey fish populations. Here, we hypothesize that European perch (Perca fluviatilis) populations being exposed to oxazepam in situ show reduced anti-predator behavior, which has previously been observed for exposed isolated fish in laboratory studies. We tested our hypothesis by exposing a whole-lake ecosystem, containing both perch (prey) and northern pike (Esox lucius; predator), to oxazepam while tracking fish behavior before and after exposure in the exposed lake as well as in an unexposed nearby lake (control). Oxazepam concentrations in the exposed lake ranged between 11 and 24 µg L-1, which is >200 times higher than concentrations reported for European rivers. In contrast to our hypothesis, we did not observe an oxazepam-induced reduction in anti-predator behavior, inferred from perch swimming activity, distance to predators, distance to conspecifics, home-range size, and habitat use. In fact, exposure to oxazepam instead stimulated anti-predator behavior (decreased activity, decreased distance to conspecifics, and increased littoral habitat use) when using behavior in the control lake as a reference. Shoal dynamics and temperature changes may have masked modest reductions in anti-predator behavior due to oxazepam. Although we cannot fully resolve the mechanism(s) behind our observations, our results indicate that the effects of oxazepam on perch behavior in a familiar natural ecosystem are negligible in comparison to the effects of other environmental conditions.

The missing pieces for better future predictions in subarctic ecosystems: A Torneträsk case study.

Arctic and subarctic ecosystems are experiencing substantial changes in hydrology, vegetation, permafrost conditions, and carbon cycling, in response to climatic change and other anthropogenic drivers, and these changes are likely to continue over this century. The total magnitude of these changes results from multiple interactions among these drivers. Field measurements can address the overall responses to different changing drivers, but are less capable of quantifying the interactions among them. Currently, a comprehensive assessment of the drivers of ecosystem changes, and the magnitude of their direct and indirect impacts on subarctic ecosystems, is missing. The Torneträsk area, in the Swedish subarctic, has an unrivalled history of environmental observation over 100 years, and is one of the most studied sites in the Arctic. In this study, we summarize and rank the drivers of ecosystem change in the Torneträsk area, and propose research priorities identified, by expert assessment, to improve predictions of ecosystem changes. The research priorities identified include understanding impacts on ecosystems brought on by altered frequency and intensity of winter warming events, evapotranspiration rates, rainfall, duration of snow cover and lake-ice, changed soil moisture, and droughts. This case study can help us understand the ongoing ecosystem changes occurring in the Torneträsk area, and contribute to improve predictions of future ecosystem changes at a larger scale. This understanding will provide the basis for the future mitigation and adaptation plans needed in a changing climate.

Decade of experimental permafrost thaw reduces turnover of young carbon and increases losses of old carbon, without affecting the net carbon balance.

Thicker snowpacks and their insulation effects cause winter-warming and invoke thaw of permafrost ecosystems. Temperature-dependent decomposition of previously frozen carbon (C) is currently considered one of the strongest feedbacks between the Arctic and the climate system, but the direction and magnitude of the net C balance remains uncertain. This is because winter effects are rarely integrated with C fluxes during the snow-free season and because predicting the net C balance from both surface processes and thawing deep layers remains challenging. In this study, we quantified changes in the long-term net C balance (net ecosystem production) in a subarctic peat plateau subjected to 10 years of experimental winter-warming. By combining 210 Pb and 14 Cdating of peat cores with peat growth models, we investigated thawing effects on year-round primary production and C losses through respiration and leaching from both shallow and deep peat layers. Winter-warming and permafrost thaw had no effect on the net C balance, but strongly affected gross C fluxes. Carbon losses through decomposition from the upper peat were reduced as thawing of permafrost induced surface subsidence and subsequent waterlogging. However, primary production was also reduced likely due to a strong decline in bryophytes cover while losses from the old C pool almost tripled, caused by the deepened active layer. Our findings highlight the need to estimate long-term responses of whole-year production and decomposition processes to thawing, both in shallow and deep soil layers, as they may contrast and lead to unexpected net effects on permafrost C storage.

Invasive earthworms unlock arctic plant nitrogen limitation.

Arctic plant growth is predominantly nitrogen (N) limited. This limitation is generally attributed to slow soil microbial processes due to low temperatures. Here, we show that arctic plant-soil N cycling is also substantially constrained by the lack of larger detritivores (earthworms) able to mineralize and physically translocate litter and soil organic matter. These new functions provided by earthworms increased shrub and grass N concentration in our common garden experiment. Earthworm activity also increased either the height or number of floral shoots, while enhancing fine root production and vegetation greenness in heath and meadow communities to a level that exceeded the inherent differences between these two common arctic plant communities. Moreover, these worming effects on plant N and greening exceeded reported effects of warming, herbivory and nutrient addition, suggesting that human spreading of earthworms may lead to substantial changes in the structure and function of arctic ecosystems.

Less anxious salmon smolt become easy prey during downstream migration.

Hatchery-reared salmon smolt used for supplementary stocking often display poor migration behavior compared to wild smolt, which reduces the success of this management action. Oxazepam, an anxiolytic drug, has been shown to intensify salmon smolt migration in mesocosm experiments, and treatment with this drug has, therefore, been suggested as a management option to improve downstream smolt migration. In this study, we tested this by assessing migration performance of hatchery-reared Atlantic salmon (Salmo salar) smolt along a 21-km long natural river-to-sea migration route in a boreal river in Northern Sweden. Using acoustic telemetry, the migration rate and survival of smolt that had been exposed to oxazepam (200 µg L-1, N = 20) was monitored and compared with a control group (N = 20) of unexposed smolt. Exposed smolt took significantly longer time to initiate migration after release compared to the control fish, but after that we observed no significant difference in downstream migration speed. However, exposed smolt had considerably higher probability of being predated on compared to control smolt. We attribute these results to increased risk-taking and higher activity in oxazepam-exposed smolt, which in turn increased initial non-directional exploratory behavior and decreased predator vigilance. These results are discussed based on current concerns for ecological implications of behavioral modifications induced by pharmaceutical pollution and climate change. We conclude that exposure to oxazepam is an unsuitable management option to prime migration of reared salmon in natural systems.

High-speed imaging reveals how antihistamine exposure affects escape behaviours in aquatic insect prey.

Aquatic systems receive a wide range of pharmaceuticals that may have adverse impacts on aquatic wildlife. Among these pharmaceuticals, antihistamines are commonly found, and these substances have the potential to influence the physiology of aquatic invertebrates. Previous studies have focused on how antihistamines may affect behaviours of aquatic invertebrates, but these studies probably do not capture the full consequences of antihistamine exposure, as traditional recording techniques do not capture important animal movements occurring at the scale of milliseconds, such as prey escape responses. In this study, we investigated if antihistamine exposure can impact escape responses in aquatic insect, by exposing damselfly (Coenagrion hastulatum) larvae to two environmentally relevant concentrations (0.1 and 1 µg L-1) of diphenhydramine. Importantly, we used a high-speed imaging approach that with high-time resolution captures details of escape responses and, thus, potential impacts of diphenhydramine on these behaviours. Our results show overall weak effects of antihistamine exposure on the escape behaviours of damselfly larvae. However, at stage 2 of the C-escape response, we found a significant increase in turning angle, which corresponds to a reduced swimming velocity, indicating a reduced success at evading a predator attack. Thus, we show that low concentrations of an antihistamine may affect behaviours strongly related to fitness of aquatic insect prey - effects that would have been overlooked using traditional recording techniques. Hence, to understand the full consequences of pharmaceutical contamination on aquatic wildlife, high-speed imaging should be incorporated into future environmental risk assessments.

Investigating tissue bioconcentration and the behavioural effects of two pharmaceutical pollutants on sea trout (Salmo trutta) in the laboratory and field.

Pharmaceuticals entering aquatic ecosystems via wastewater effluents are of increasing concern for wild animals. Because some pharmaceuticals are designed to modulate human behaviour, measuring the impacts of exposure to pharmaceuticals on fish behaviour has become a valuable endpoint. While laboratory studies have shown that pharmaceuticals can affect fish behaviour, there is a lack of understanding if behaviour is similarly affected in natural environments. Here, we exposed sea trout (Salmo trutta) smolts to two concentrations of two pharmaceutical pollutants often detected in surface waters: temazepam (a benzodiazepine, anxiolytic) or irbesartan (an angiotensin II receptor blocker, anti-hypertensive). We tested the hypothesis that changes to behavioural traits (anxiety and activity) measured in laboratory trials following exposure are predictive of behaviour in the natural environment (downstream migration). Measures of anxiety and activity in the laboratory assay did not vary with temazepam treatment, but temazepam-exposed fish began migrating faster in the field. Activity in the laboratory assay did predict overall migration speed in the field. In contrast to temazepam, we found that irbesartan exposure did not affect behaviour in the laboratory, field, or the relationship between the two endpoints. However, irbesartan was also not readily taken up into fish tissue (i.e. below detection levels in the muscle tissue), while temazepam bioconcentrated (bioconcentration factor 7.68) rapidly (t1/2 < 24 h). Our findings add to a growing literature showing that benzodiazepine pollutants can modulate fish behaviour and that laboratory assays may be less sensitive at detecting the effects of pollutants compared to measuring effects in natural settings. Therefore, we underscore the importance of measuring behavioural effects in the natural environment.

Behavioural effects of psychoactive pharmaceutical exposure on European perch (Perca fluviatilis) in a multi-stressor environment.

With the ability to resist biodegradation and exert therapeutic effects at low concentrations, pharmaceutical contaminants have become environmental stressors for wildlife. One such contaminant is the anxiolytic oxazepam, a psychoactive pharmaceutical that is frequently detected in surface waters globally. Despite growing interest in understanding how wildlife respond to anxiolytics, synergistic effects of pharmaceuticals and other abiotic (e.g. temperature) and biotic (e.g. predation risk) stressors remain unclear. Here, using a multi-stressor approach, we investigated effects of 7-day oxazepam exposure (6.5 µg/L) on anxiety-related behaviours in juvenile European perch (Perca fluviatilis). The multi-stressor approach was achieved by exposing perch to oxazepam at two temperatures (10 °C and 18 °C), and at two predation risk regimes-generated using chemical cues from the northern pike (Esox lucius). Our exposures resulted in a successful uptake of the drug from the water, i.e., oxazepam was measured in perch muscle tissue at 50 ±â€¯17 ng/g (mean ±â€¯SD). We found significant oxazepam-induced effects on boldness, with 76.7% of the treated fish entering the white background (i.e. 'exposed' area where exposure to presumed risks are higher) within the first 5 min, compared to 66.6% of the control fish. We also found a significant effect of temperature on total time spent freezing (i.e. staying motionless). Specifically, fish in the low temperature treatments (oxazepam, predation) froze for longer than fish in high temperatures. Our multi-stressor study is the first to uncover how anxiety-related behaviours in wild juvenile fish are altered by changes in water temperature and perceived predation risk. Importantly, our findings highlight the need to focus on multiple stressors to improve understanding of how organisms not only survive, but adapt to, human-induced environmental change.

ToxId: an efficient algorithm to solve occlusions when tracking multiple animals.

Video analysis of animal behaviour is widely used in fields such as ecology, ecotoxicology, and evolutionary research. However, when tracking multiple animals, occlusion and crossing are problematic, especially when the identity of each individual needs to be preserved. We present a new algorithm, ToxId, which preserves the identity of multiple animals by linking trajectory segments using their intensity histogram and Hu-moments. We verify the performance and accuracy of our algorithm using video sequences with different animals and experimental conditions. The results show that our algorithm achieves state-of-the-art accuracy using an efficient approach without the need of learning processes, complex feature maps or knowledge of the animal shape. ToxId is also computationally efficient, has low memory requirements, and operates without accessing future or past frames.

Environmental relevant levels of a benzodiazepine (oxazepam) alters important behavioral traits in a common planktivorous fish, (Rutilus rutilus).

Environmental pollution by pharmaceuticals is increasingly recognized as a major threat to aquatic ecosystems worldwide. A complex mix of pharmaceuticals enters waterways via treated wastewater effluent and many remain biochemically active after the drugs reach aquatic systems. However, to date little is known regarding the ecological effects that might arise following pharmaceutical contamination of aquatic environments. One group of particular concern is behaviorally modifying pharmaceuticals as seemingly minor changes in behavior may initiate marked ecological consequences. The aim of this study was to examine the influence of a benzodiazepine anxiolytic drug (oxazepam) on key behavioral traits in wild roach (Rutilus rutilus) at concentrations similar to those encountered in effluent surface waters. Roach exposed to water with high concentrations of oxazepam (280 µg/L) exhibited increased boldness, while roach at low treatment (0.84 µg/L) became bolder and more active compared to control fish. Our results reinforce the notion that anxiolytic drugs may be affecting fish behavior in natural systems, emphasizing the need for further research on ecological impacts of pharmaceuticals in aquatic systems and development of new tools to incorporate ecologically relevant behavioral endpoints into ecotoxicological risk assessment.

Refining particle positions using circular symmetry.

Particle and object tracking is gaining attention in industrial applications and is commonly applied in: colloidal, biophysical, ecological, and micro-fluidic research. Reliable tracking information is heavily dependent on the system under study and algorithms that correctly determine particle position between images. However, in a real environmental context with the presence of noise including particular or dissolved matter in water, and low and fluctuating light conditions, many algorithms fail to obtain reliable information. We propose a new algorithm, the Circular Symmetry algorithm (C-Sym), for detecting the position of a circular particle with high accuracy and precision in noisy conditions. The algorithm takes advantage of the spatial symmetry of the particle allowing for subpixel accuracy. We compare the proposed algorithm with four different methods using both synthetic and experimental datasets. The results show that C-Sym is the most accurate and precise algorithm when tracking micro-particles in all tested conditions and it has the potential for use in applications including tracking biota in their environment.

Screening of benzodiazepines in thirty European rivers.

Pharmaceuticals as environmental contaminants have received a lot of interest over the past decade but, for several pharmaceuticals, relatively little is known about their occurrence in European surface waters. Benzodiazepines, a class of pharmaceuticals with anxiolytic properties, have received interest due to their behavioral modifying effect on exposed biota. In this study, our results show the presence of one or more benzodiazepine(s) in 86% of the analyzed surface water samples (n = 138) from 30 rivers, representing seven larger European catchments. Of the 13 benzodiazepines included in the study, we detected 9, which together showed median and mean concentrations (of the results above limit of quantification) of 5.4 and 9.6 ng L-1, respectively. Four benzodiazepines (oxazepam, temazepam, clobazam, and bromazepam) were the most commonly detected. In particular, oxazepam had the highest frequency of detection (85%) and a maximum concentration of 61 ng L-1. Temazepam and clobazam were found in 26% (maximum concentration of 39 ng L-1) and 14% (maximum concentration of 11 ng L-1) of the samples analyzed, respectively. Finally, bromazepam was found only in Germany and in 16 out of total 138 samples (12%), with a maximum concentration of 320 ng L-1. This study clearly shows that benzodiazepines are common micro-contaminants of the largest European river systems at ng L-1 levels. Although these concentrations are more than a magnitude lower than those reported to have effective effects on exposed biota, environmental effects cannot be excluded considering the possibility of additive and sub-lethal effects.

Boreal Forests Sequester Large Amounts of Mercury over Millennial Time Scales in the Absence of Wildfire.

Alterations in fire activity due to climate change and fire suppression may have profound effects on the balance between storage and release of carbon (C) and associated volatile elements. Stored soil mercury (Hg) is known to volatilize due to wildfires and this could substantially affect the land-air exchange of Hg; conversely the absence of fires and human disturbance may increase the time period over which Hg is sequestered. Here we show for a wildfire chronosequence spanning over more than 5000 years in boreal forest in northern Sweden that belowground inventories of total Hg are strongly related to soil humus C accumulation (R2 = 0.94, p < 0.001). Our data clearly show that northern boreal forest soils have a strong sink capacity for Hg, and indicate that the sequestered Hg is bound in soil organic matter pools accumulating over millennia. Our results also suggest that more than half of the Hg stock in the sites with the longest time since fire originates from deposition predating the onset of large-scale anthropogenic emissions. This study emphasizes the importance of boreal forest humus soils for Hg storage and reveals that this pool is likely to persist over millennial time scales in the prolonged absence of fire.

GABAergic anxiolytic drug in water increases migration behaviour in salmon.

Migration is an important life-history event in a wide range of taxa, yet many migrations are influenced by anthropogenic change. Although migration dynamics are extensively studied, the potential effects of environmental contaminants on migratory physiology are poorly understood. In this study we show that an anxiolytic drug in water can promote downward migratory behaviour of Atlantic salmon (Salmo salar) in both laboratory setting and in a natural river tributary. Exposing salmon smolt to a dilute concentration of a GABAA receptor agonist (oxazepam) increased migration intensity compared with untreated smolt. These results implicate that salmon migration may be affected by human-induced changes in water chemical properties, such as acidification and pharmaceutical residues in wastewater effluent, via alterations in the GABAA receptor function.