Nanoplastic pollution changes the intestinal microbiome but not the morphology or behavior of a freshwater turtle.

Humans produce 350 million metric tons of plastic waste per year, leading to microplastic pollution and widespread environmental contamination, particularly in aquatic environments. This subsequently impacts aquatic organisms in myriad ways, yet the vast majority of research is conducted in marine, rather than freshwater systems. In this study, we exposed eggs and hatchlings of the Chinese soft-shelled turtle (Pelodiscus sinensis) to 80-nm polystyrene nanoplastics (PS-NPs) and monitored the impacts on development, behavior and the gut microbiome. We demonstrate that 80-nm PS-NPs can penetrate the eggshell and move into developing embryos. This led to metabolic impairments, as evidenced by bradycardia (a decreased heart rate), which persisted until hatching. We found no evidence that nanoplastic exposure affected hatchling morphology, growth rates, or levels of boldness and exploration, yet we discuss some potential caveats here. Exposure to nanoplastics reduced the diversity and homogeneity of gut microbiota in P. sinensis, with the level of disruption correlating to the length of environmental exposure (during incubation only or post-hatching also). Thirteen core genera (with an initial abundance >1 %) shifted after nanoplastic treatment: pathogenic bacteria increased, beneficial probiotic bacteria decreased, and there was an increase in the proportion of negative correlations between bacterial genera. These changes could have profound impacts on the viability of turtles throughout their lives. Our study highlights the toxicity of environmental NPs to the embryonic development and survival of freshwater turtles. We provide insights about population trends of P. sinensis in the wild, and future directions for research.

The ethics of intervening in animal behaviour for conservation.

Conservation behaviour is a growing field that applies insights from the study of animal behaviour to address challenges in wildlife conservation and management. Conservation behaviour interventions often aim to manage specific behaviours of a species to solve conservation challenges. The field is often viewed as offering approaches that are less intrusive or harmful to animals than, for example, managing the impact of a problematic species by reducing its population size (frequently through lethal control). However, intervening in animal behaviour, even for conservation purposes, may still raise important ethical considerations. We discuss these issues and develop a framework and a decision support tool, to aid managers and researchers in evaluating the ethical considerations of conservation behaviour interventions against other options.

Taking the bait: Developing a bait delivery system to target free-ranging crocodiles and varanid lizards with a novel conservation strategy.

In tropical Australia, conditioned taste aversion (CTA) can buffer vulnerable native predators from the invasion of a toxic prey species (cane toads, Rhinella marina). Thus, we need to develop methods to deploy aversion-inducing baits in the field, in ways that maximize uptake by vulnerable species (but not other taxa). We constructed and field-tested baiting devices, in situ with wild animals. Apparatus were set next to waterbodies and baited concurrently at multiple locations (over water, water's edge, and on the bank). Baits were checked and replaced twice daily during the trial; remote cameras recorded visitation by native predators. Bait longevity was compared at sun-exposed and shaded locations over 12 h. The strength required to remove baits from apparatus was measured in varanids and crocodiles. The device promoted high rates of bait uptake by freshwater crocodiles (47% baits consumed), varanid lizards (19% baits consumed), and non-target taxa (34% baits consumed). Targeting specific predators can be achieved by manipulating bait location and time of deployment, as well as the force required to dislodge the bait. Crocodiles were best targeted with over-water baits, whereas varanid lizards preferred baits located at the edges of waterbodies. When testing bait longevity in ambient conditions, during the daytime baits desiccated fully within 12 h, and faster in the sun than in the shade. Based on studies using captive animals, the "pulling force" strength of reptilian predators scaled with body size and was greater in crocodiles than in varanid lizards. We present the first conservation baiting protocol designed specifically for reptiles. Our results demonstrate the feasibility of widespread and taxon-specific deployment of aversion-inducing baits to buffer the impacts of invasive cane toads, and our methods are applicable (with modification) to other research and management programs globally.

Shifts in the foraging tactics of crocodiles following invasion by toxic prey.

Biological invasions can modify the behaviour of vulnerable native species in subtle ways. For example, native predators may learn or evolve to reduce foraging in conditions (habitats, times of day) that expose them to a toxic invasive species. In tropical Australia, freshwater crocodiles (Crocodylus johnstoni) are often fatally poisoned when they ingest invasive cane toads (Rhinella marina). The risk may be greatest if toads are seized on land, where a predator cannot wash away the toxins before they are absorbed into its bloodstream. Hence, toad invasion might induce crocodiles to forage in aquatic habitats only, foregoing terrestrial hunting. To test this idea, we conducted standardised trials of bait presentation to free-ranging crocodiles in sites with and without invasive toads. As anticipated, crocodiles rapidly learned to avoid consuming toads, and shifted to almost exclusively aquatic foraging.

Increased rates of dispersal of free-ranging cane toads (Rhinella marina) during their global invasion.

Invasions often accelerate through time, as dispersal-enhancing traits accumulate at the expanding range edge. How does the dispersal behaviour of individual organisms shift to increase rates of population spread? We collate data from 44 radio-tracking studies (in total, of 650 animals) of cane toads (Rhinella marina) to quantify distances moved per day, and the frequency of displacement in their native range (French Guiana) and two invaded areas (Hawai'i and Australia). We show that toads in their native-range, Hawai'i and eastern Australia are relatively sedentary, while toads dispersing across tropical Australia increased their daily distances travelled from 20 to 200 m per day. That increase reflects an increasing propensity to change diurnal retreat sites every day, as well as to move further during each nocturnal displacement. Daily changes in retreat site evolved earlier than did changes in distances moved per night, indicating a breakdown in philopatry before other movement behaviours were optimised to maximise dispersal.

Divergent long-term impacts of lethally toxic cane toads (Rhinella marina) on two species of apex predators (monitor lizards, Varanus spp.).

Biological invasions can massively disrupt ecosystems, but evolutionary and ecological adjustments may modify the magnitude of that impact through time. Such post-colonisation shifts can change priorities for management. We quantified the abundance of two species of giant monitor lizards, and of the availability of their mammalian prey, across 45 sites distributed across the entire invasion trajectory of the cane toad (Rhinella marina) in Australia. One varanid species (Varanus panoptes from tropical Australia) showed dramatic population collapse with toad invasion, with no sign of recovery at most (but not all) sites that toads had occupied for up to 80 years. In contrast, abundance of the other species (Varanus varius from eastern-coastal Australia) was largely unaffected by toad invasion. That difference might reflect availability of alternative food sources in eastern-coastal areas, perhaps exacerbated by the widespread prior collapse of populations of small mammals across tropical (but not eastern) Australia. According to this hypothesis, the impact of cane toads on apex predators has been exacerbated and prolonged by a scarcity of alternative prey. More generally, multiple anthropogenically-induced changes to natural ecosystems may have synergistic effects, intensifying the impacts beyond that expected from either threat in isolation.

Untangling the influence of biotic and abiotic factors on habitat selection by a tropical rodent.

Understanding how animal populations respond to environmental factors is critical because large-scale environmental processes (e.g., habitat fragmentation, climate change) are impacting ecosystems at unprecedented rates. On an overgrazed floodplain in north-western Australia, a native rodent (Pale Field Rat, Rattus tunneyi) constructs its burrows primarily beneath an invasive tree (Chinee Apple, Ziziphus mauritiana) rather than native trees. The dense thorny foliage of the Chinee Apple may allow high rat densities either because of abiotic effects (shade, in a very hot environment) or biotic processes (protection from trampling and soil compaction by feral horses, and/or predation). To distinguish between these hypotheses, we manipulated Chinee Apple foliage to modify biotic factors (access to horses and predators) but not shade levels. We surveyed the rat population with Elliott traps under treatment and control trees and in the open woodland, in two seasons (the breeding season-January, and the nesting season-May). In the breeding season, we ran giving-up density experiments (GUD) with food trays, to assess the perceived risk of predation by rats across our three treatments. Selective trimming of foliage did not affect thermal regimes underneath the trees but did allow ingress of horses and we observed two collapsed burrows as a consequence (although long term impacts of horses were not measured). The perceived predation risk also increased (GUD values at food trays increased) and was highest in the open woodland. Our manipulation resulted in a shift in rat sex ratios (indicating female preference for breeding under control but not foliage-trimmed trees) and influenced rat behaviour (giving-up densities increased; large dominant males inhabited the control but not treatment trees). Our data suggest that the primary benefit of the Chinee Apple tree to native rodents lies in physical protection from predators and (potentially) feral horses, rather than in providing cooler microhabitat.

Characterization of the complete mitochondrial genome of the many-lined sun skink (Eutropis multifasciata) and comparison with other Scincomorpha species.

Characterizating the complete mitochondrial genome (mitogenome) of an organism allows detailed genomic studies in systematics and evolution. The present study decodes the mitogenome (17,062 bp) of the many-lined sun skink, Eutropis multifasciata, using next-generation sequencing. To compare the diversity of mitogenomic structure and investigate intraspecific evolutionary relationships among the Asian Scincomorpha, the mitogenomes of 46 other species were examined concurrently. Within the group, the size of mitogenomes varied predominantly in the length at their control regions. The Ka/Ks ratios of 12 protein codon genes (PCGs) were lower than 1.00, demonstrating that they were under relaxed or moderate purifying selection. However, the ND5 had a Ka/Ks ratio >1, and was considered to be under positive selection. Currently there are two superfamilies in Scincomorpha (i.e. Scincoidea and Lacertoidea), but phylogenetic analysis using Bayesian Inference and Maximum-Likelihood Estimations produced phylogenetic trees with three clades in Scincomorpha ((Scincoidea + Lacertoidea (part)) + Gymnophthalmidae)).

Anthropogenically modified habitats favor bigger and bolder lizards.

Anthropogenic activities often create distinctive but discontinuously distributed habitat patches with abundant food but high risk of predation. Such sites can be most effectively utilized by individuals with specific behaviors and morphologies. Thus, a widespread species that contains a diversity of sizes and behavioral types may be pre-adapted to exploiting such hotspots. In eastern Australia, the giant (to >2 m) lizard Varanus varius (lace monitor) utilizes both disturbed (campground) and undisturbed (bushland) habitats. Our surveys of 27 sites show that lizards found in campgrounds tended to be larger and bolder than those in adjacent bushland. This divergence became even more marked after the arrival of a toxic invasive species (the cane toad, Rhinella marina) caused high mortality in larger and bolder lizards. Some of the behavioral divergences between campground and bushland lizards may be secondary consequences of differences in body size, but other habitat-associated divergences in behavior are due to habituation and/or nonrandom mortality.

Choose your meals carefully if you need to coexist with a toxic invader.

Vulnerable native species may survive the impact of a lethally toxic invader by changes in behaviour, physiology and/or morphology. The roles of such mechanisms can be clarified by standardised testing. We recorded behavioural responses of monitor lizards (Varanus panoptes and V. varius) to legs of poisonous cane toads (Rhinella marina) and non-toxic control meals (chicken necks or chicken eggs and sardines) along 1300 and 2500 km transects, encompassing the toad's 85-year invasion trajectory across Australia as well as yet-to-be-invaded sites to the west and south of the currently colonised area. Patterns were identical in the two varanid species. Of monitors that consumed at least one prey type, 96% took control baits whereas toad legs were eaten by 60% of lizards in toad-free sites but 0% from toad-invaded sites. Our survey confirms that the ability to recognise and reject toads as prey enables monitor lizards to coexist with cane toads. As toxic invaders continue to impact ecosystems globally, it is vital to understand the mechanisms that allow some taxa to persist over long time-scales.

Diurnal activity in cane toads (Rhinella marina) is geographically widespread.

Although adult cane toads (Rhinella marina) are generally active only at night, a recent study reported that individuals of this species switched to diurnal activity in response to encountering a novel habitat type (deeply shaded gorges) in the course of their Australian invasion. Our sampling over a broader geographic scale challenges the idea that this behaviour is novel; we documented diurnal behaviour both in the species' native range and in several sites within the invaded range, in multiple habitat types. Diurnal activity was most common in the tropics and in areas where toads attain high population densities and are in poor body condition, suggesting that the expansion of activity times may be induced by intraspecific competition for food.

Authorship Protocols Must Change to Credit Citizen Scientists.

The sociopolitical nature of research is changing and so must our protocols for authorship. Citizen scientists are often excluded from authorship because they cannot meet rigid journal criteria. To address this, we propose a new concept: allowing nonprofessional scientists to be credited as authors under a collective identity ('group coauthorship').

New Weapons in the Toad Toolkit: A Review of Methods to Control and Mitigate the Biodiversity Impacts of Invasive Cane Toads (Rhinella Marina).

Our best hope of developing innovative methods to combat invasive species is likely to come from the study of high-profile invaders that have attracted intensive research not only into control, but also basic biology. Here we illustrate that point by reviewing current thinking about novel ways to control one of the world's most well-studied invasions: that of the cane toad in Australia. Recently developed methods for population suppression include more effective traps based on the toad's acoustic and pheromonal biology. New tools for containing spread include surveillance technologies (e.g., eDNA sampling and automated call detectors), as well as landscape-level barriers that exploit the toad's vulnerability to desiccation­a strategy that could be significantly enhanced through the introduction of sedentary, range-core genotypes ahead of the invasion front. New methods to reduce the ecological impacts of toads include conditioned taste aversion in free-ranging predators, gene banking, and targeted gene flow. Lastly, recent advances in gene editing and gene drive technology hold the promise of modifying toad phenotypes in ways that may facilitate control or buffer impact. Synergies between these approaches hold great promise for novel and more effective means to combat the toad invasion and its consequent impacts on biodiversity.

Eliciting conditioned taste aversion in lizards: Live toxic prey are more effective than scent and taste cues alone.

Conditioned taste aversion (CTA) is an adaptive learning mechanism whereby a consumer associates the taste of a certain food with symptoms caused by a toxic substance, and thereafter avoids eating that type of food. Recently, wildlife researchers have employed CTA to discourage native fauna from ingesting toxic cane toads (Rhinella marina), a species that is invading tropical Australia. In this paper, we compare the results of 2 sets of CTA trials on large varanid lizards ("goannas," Varanus panoptes). One set of trials (described in this paper) exposed recently-captured lizards to sausages made from cane toad flesh, laced with a nausea-inducing chemical (lithium chloride) to reinforce the aversion response. The other trials (in a recently-published paper, reviewed herein) exposed free-ranging lizards to live juvenile cane toads. The effectiveness of the training was judged by how long a lizard survived in the wild before it was killed (fatally poisoned) by a cane toad. Both stimuli elicited rapid aversion to live toads, but the CTA response did not enhance survival rates of the sausage-trained goannas after they were released into the wild. In contrast, the goannas exposed to live juvenile toads exhibited higher long-term survival rates than did untrained conspecifics. Our results suggest that although it is relatively easy to elicit short-term aversion to toad cues in goannas, a biologically realistic stimulus (live toads, encountered by free-ranging predators) is most effective at buffering these reptiles from the impact of invasive toxic prey.

Toads on Lava: Spatial Ecology and Habitat Use of Invasive Cane Toads (Rhinella marina) in Hawai'i.

Most ecological research on cane toads (Rhinella marina) has focused on invasive populations in Australia, ignoring other areas where toads have been introduced. We radio-tracked and spool-tracked 40 toads, from four populations on the island of Hawai'i. Toads moved extensively at night (mean 116 m, from spool-tracking) but returned to the same or a nearby retreat-site each day (from radio-tracking, mean distance between successive retreat sites 11 m; 0 m for 70% of records). Males followed straighter paths during nocturnal movements than did females. Because moist sites are scarce on the highly porous lava substrate, Hawai'ian toads depend on anthropogenic disturbance for shelter (e.g. beneath buildings), foraging (e.g. suburban lawns, golf courses) and breeding (artificial ponds). Foraging sites are further concentrated by a scarcity of flying insects (negating artificial lights as prey-attractors). Habitat use of toads shifted with time (at night, toads selected areas with less bare ground, canopy, understory and leaf-litter), and differed between sexes (females foraged in areas of bare ground with dense understory vegetation). Cane toads in Hawai'i thrive in scattered moist patches within a severely arid matrix, despite a scarcity of flying insects, testifying to the species' ability to exploit anthropogenic disturbance.