Léa Fieschi-Méric

In the midst of the current biodiversity crisis, amphibians are severely threatened by emerging diseases such as chytridiomycosis. Characterizing the mechanisms that underlie susceptibility to this disease is fundamental to improve amphibian conservation. Using a comprehensive multi‐omics approach, this study investigates the impact of an exposure to the chytrid fungus Batrachochytrium salamandrivorans ( Bsal ) on the gene expression of Italian crested newts ( Triturus carnifex ) and on the bacterial symbionts that constitute their skin microbiota. Exposure to Bsal affected multiple components of the newts' immunity, from limited structural changes in their microbiota and decreased expression of keratin‐encoding genes in the skin to the upregulation of genes involved in inflammation and adaptive immunity both at the site of infection (skin) and in their primary lymphoid organ (spleen). We found that chytridiomycosis severity was positively correlated with the downregulation of basal metabolism and with the upregulation of immune responses and of tissue restructuration. Together, our results suggest that in T. carnifex , Bsal susceptibility may be linked to a reallocation of energy resources from the maintenance of basal metabolism and tissue integrity towards elevated immunity and tissue restructuration.

The European hedgehog ( Erinaceus europaeus ) is currently designated as near threatened by the International Union for the Conservation of Nature. Threats include reduced prey availability, emerging pathogens, and environmental contaminants. Improving knowledge of food preferences and the role of diet in disease and exposure to toxins is important for its conservation. We used fecal DNA metabarcoding to characterize the diet of 209 hedgehogs in Europe. Stylommatophora (slugs and snails) made up the most frequent order in the hedgehog diet (found in 92% of individuals), followed by Haplotaxida (earthworms; 54%), Coleoptera (beetles; 54%), Lepidoptera (moths and butterflies; 48%), and Orthoptera (grasshoppers, locusts, and crickets; 54%). Presumed anthropogenic food, primarily in the form of pet food, was consumed by 29% of hedgehogs. We found no relation between diet and infection with pathogens ( Corynebacterium ulcerans or circovirus). However, diet richness was negatively correlated with hepatic concentration in brodifacoum, a widely used rodenticide. Moreover, the relative abundance of Stylommatophora in the hedgehog diet was positively correlated with their hepatic concentration of zinc. Generally, Stylommatophora consumption was most frequent at locations of highest cumulated rainfall. The near‐ubiquity of invasive Spanish slugs ( Arion vulgaris ; found in 80% of individuals) in the diet is concerning for hedgehog conservation, since this species is a known vector of lungworm parasites and accumulates high concentrations of environmental contaminants, such as zinc and rodenticides. Overall, this study provides a novel, high‐resolution view of hedgehog diet composition and highlights the need for further research on the role of terrestrial gastropods as toxicant vectors through food webs.

Background: Amidst the current biodiversity crisis, amphibians are particularly endangered by the emergence of infectious diseases. The skin disease chytridiomycosis is caused by the fungi Batrachochytrium dendrobatidis (Bd) and B. salamandrivorans (Bsal) , which may interact with bacterial symbionts present on the amphibian epidermis. Extensive research has explored the interactions between the amphibian microbiota and Bd ; yet, little is known about its interactions with Bsal . In this paper, we used the ribbed newt ( Pleurodeles waltl ), a model species displaying pronounced among-individual variation in response to Bsal, to (1) determine whether susceptibility to Bsal and individual microbiota vary between source groups; (2) test whether susceptibility to Bsal can be predicted from skin microbiota before exposure and (3) quantify microbiota volatility over time to determine whether Bsal infection intensity and chytridiomycosis severity correlate with the magnitude of shifts in bacterial communities caused by Bsal exposure. Results: Our results demonstrate that newts of different origin harbor distinct microbiota even under uniform rearing conditions. We show that Bsal infection intensity and disease severity cannot be predicted from the diversity, structure, or composition of the skin microbiota of P. waltl. Instead, a strong relation between newts’ source group and their response to Bsal suggests that other factors might underpin among-individual variation in Bsal susceptibility in this species. Moreover, our results indicate that the intensity of early Bsal infection and longer-term severity of chytridiomycosis do not correlate with the magnitude of microbiota change following Bsal exposure. Conclusion: These results demonstrate a limited involvement of the microbiota in Bsal dynamics in P. waltl, suggesting that other mechanisms contribute to individual Bsal susceptibility. Further research on the relation between chytrid pathogens and their amphibian hosts will be instrumental to improve the conservation of the most endangered vertebrate class on earth. Graphical

Amphibians face a multitude of threats and therefore make a prime example of the current biodiversity crisis. Multiple amphibian stressors in anthropogenic landscapes include infectious diseases and agrochemicals. Synergic effects between these stressors may increase the negative impact of infections on amphibian health. In a 56-day trial, we assessed the impact of co-exposure to the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) and the pathogenic fungus Batrachochytrium salamandrivorans (Bsal) on infection parameters (infection intensity and disease severity) and health (body condition and telomere length) in Italian crested newts (Triturus carnifex). Twenty days post-inoculation with Bsal, newts were either exposed to 2,4-D for 12 days or not exposed (control). Most newts developed high infection loads that steadily increased towards the end of the trial. While body condition was negatively correlated with pathogen burden, only one out of 23 animals died. Telomere length remained unaffected by the pesticide and the pathogen. The 2,4-D treatment did not exacerbate Bsal infection. Most newts survived almost two months with significant pathogen loads; thus, even in a pesticide-infested environment, T. carnifex could be an important long-term Bsal reservoir for co-occurring species on the Italian peninsula, a urodele diversity hotspot.

The key role of symbiotic skin bacteria communities in amphibian resistance to emerging pathogens is well recognized, but factors leading to their dysbiosis are not fully understood. In particular, the potential effects of population translocations on the composition and diversity of hosts' skin microbiota have received little attention, although such transfers are widely carried out as a strategy for amphibian conservation. To characterize the potential reorganization of the microbiota over such a sudden environmental change, we conducted a common‐garden experiment simulating reciprocal translocations of yellow‐spotted salamander larvae across three lakes. We sequenced skin microbiota samples collected before and 15 days after the transfer. Using a database of antifungal isolates, we identified symbionts with known function against the pathogen Batrachochytrium dendrobatidis, a major driver of amphibian declines. Our results indicate an important reorganization of bacterial assemblages throughout ontogeny, with strong changes in composition, diversity and structure of the skin microbiota in both control and translocated individuals over the 15 days of monitoring. Unexpectedly, the diversity and community structure of the microbiota were not significantly affected by the translocation event, thus suggesting a strong resilience of skin bacterial communities to environmental change—at least across the time‐window studied here. A few phylotypes were more abundant in the microbiota of translocated larvae, but no differences were found among pathogen‐inhibiting symbionts. Taken together, our results support amphibian translocations as a promising strategy for this endangered animal class, with limited impact on their skin microbiota.

In response to the current worldwide amphibian extinction crisis, conservation instances have encouraged the establishment of ex-situ collections for endangered species. The resulting assurance populations are managed under strict biosecure protocols, often involving artificial cycles of temperature and humidity to induce active and overwintering phases, which likely affect the bacterial symbionts living on the amphibian skin. However, the skin microbiota is an important first line of defense against pathogens that can cause amphibian declines, such as the chytrid Batrachochytrium dendrobatidis (Bd). Determining whether current husbandry practices for assurance populations might deplete amphibians from their symbionts is therefore essential to conservation success. Here, we characterize the effect of the transitions from the wild to captivity, and between aquatic and overwintering phases, on the skin microbiota of two newt species. While our results confirm differential selectivity of skin microbiota between species, they underscore that captivity and phase-shifts similarly affect their community structure. More specifically, the translocation ex-situ is associated with rapid impoverishment, decrease in alpha diversity and strong species turnover of bacterial communities. Shifts between active and overwintering phases also cause changes in the diversity and composition of the microbiota, and on the prevalence of Bd-inhibitory phylotypes. Altogether, our results suggest that current husbandry practices strongly restructure the amphibian skin microbiota. Although it remains to be determined whether these changes are reversible or have deleterious effects on their hosts, we discuss methods to limit microbial diversity loss ex-situ and emphasize the importance of integrating bacterial communities to applied amphibian conservation.

The interest in the welfare of zoo animals, from both the public and the scientific community, has long been biased towards mammals. However, growing evidence of the complex behavioural repertoires of less charismatic animals, such as reptiles, reveals the necessity to better comply with their welfare needs in captivity. Here, we present the effects of an enclosure change towards a more natural habitat in captive Galapagos tortoises (Chelonoidis spp.) held at ZSL London Zoo. Using behavioural observations, we found that the tortoises habituated to their new enclosure in six days. This represents the first quantification of habituation latency to a new enclosure in a reptile model to our knowledge—which is important information to adapt policies governing animal moves. The tortoises expressed time budgets more similar to those of wild individuals after their transition to the new enclosure. Interestingly, the hierarchy between the individuals was inverted and more stable after this change in environment. The tortoises interacted less often, which led to a decrease in the frequency of agonistic encounters. We also found that higher ambient sound volume was associated with increased likelihood of interactions turning into fights. Taken together, our results demonstrate the potential of appropriate enclosure design to improve reptile welfare.

Maintaining Gymnophiona in captivity provides opportunities to study the behaviour and life-history of this poorly known Order, and to investigate and provide species-appropriate welfare guidelines, which are currently lacking. This study focuses on the terrestrial caecilian Herpele squalostoma to investigate its sensitivity to disturbances associated with routine husbandry needed for monitoring and maintaining adequate wellbeing in captivity. Fossorial caecilians gradually pollute their environment in captivity with waste products, and substrate must be replaced at intervals; doing so disturbs the animals directly and via destruction of burrow networks. As inappetence is frequently associated with stress in amphibians, the percentage consumption of offered food types, river shrimp (Palaemon varians) and brown crickets (Gryllus assimilis), was measured as an indicator of putative stress following three routine substrate changes up to 297 days post-substrate change. Mean daily variation in substrate temperatures were also recorded in order to account for environmental influences on food consumption, along with nitrogenous waste in tank substrate prior to a substrate change and fresh top soil in order to understand the trade-off between dealing with waste accumulation and disturbing animals. We found a significant negative effect of substrate disturbance on food intake, but no significant effect of prey type. Variations in daily soil temperatures did not have a significant effect on food intake, but mean substrate temperature did. Additionally, substrate nitrogenous waste testing indicated little difference between fresh and tank substrate. In conclusion, this study provides a basis from which to develop further welfare assessment for this and other rarely kept and rarely observed terrestrial caecilian species.

Visitors to zoos can have positive, neutral, or negative relationships with zoo animals. This makes human–animal interactions (HAIs) an essential component of welfare and an important consideration in species selection for zoo exhibits and in enclosure designs. We measured the effect of visitors on reptiles by comparing open and closed periods during the lockdowns in response to the COVID-19 pandemic in the UK in a low-resolution dataset for thirteen species of reptiles and a high-resolution dataset focussing on just one of these. Scan sampling on thirteen reptile species (two chelonians and eleven squamates) showed species-specific differences in response to the presence/absence of visitors, with most taxa being only weakly affected. High-resolution scan sampling via video footage of an off-show and on-show enclosure was carried out for tokay geckos (Gekko gecko) over the open and closed periods. In this part of the study, tokay geckos were significantly more visible during zoo closure than when visitors were present on-exhibit, but there was no change in off-show animals, indicating the effect of visitors as opposed to other factors, such as seasonality, which applied equally to both on- and off-show animals. The high-resolution study showed that a significant effect was present for tokay geckos, even though the low-resolution suggested that they were more weakly affected than other taxa. Our results indicate that, for cryptic species such as this, more intensive sampling may be required to properly understand visitor effects. Our data do not allow the interpretation of effects on welfare but show that such assessments require a species-specific approach.