When carapace governs size: variation among age classes and individuals in a free-ranging ectotherm with delayed maturity.

Juvenile growth strongly impacts life-history traits during adulthood. Yet, in juveniles with delayed maturity, elusiveness has hindered age-specific studies of growth, precluding any detailed research on its consequences later in life. Different complex growth patterns have been extracted from captive animals, suggesting species-specific trajectories occur in free-ranging animals. How pronounced are growth and body size variation (VBS) throughout a long-lived ectotherm's life? Is VBS constant among age classes prior to maturity, or do compensatory and/or cumulative effects driven by long-lived-animal-specific strategies create distinct VBS cohorts, to ensure survival? To tackle the issue, we modelled growth data from continuous and dense annual capture-mark-recapture sampling (5096 body measurements of 1134 free-ranging individuals) of both immature and mature, long-lived Hermann's tortoises. We analysed population, cohort, and individual-based growth and VBS. Growth ring inferred ages were cross validated with annual recaptures in 289 juveniles. Analyses unravelled an S-shaped growth curve and identified three age cohorts across which VBS increases in a step-wise manner. Neonate-specific constraints and compensatory effects seem to control VBS until 4 years of age, possibly promoting survival with size. Subsequently, a hardened carapace takes over and cumulative effects fuelled by faster growth progressively increase VBS. Whereas ungulates are in a hurry to attain adult size before growth ceases (minimizing VBS), indeterminately growing tortoises can shape individual asymptotic sizes even after growth decelerates. Tortoise size is clearly shaped by age-specific ecological constraints; interestingly, it is likely the carapace that conducts the strategy, rather than maturity per se.

Body condition scoring facilitates healthcare monitoring in Hermann's Tortoises (Testudo hermanni ssp.).

Clinical assessment of body condition is crucial in captive and free-ranging reptiles, since a large percentage of diseases result from inadequate nutrition. However, preventive health care is restricted by the lack of a practical method for the assessment in tortoises. Pre-existing evaluation systems based on weight and shell measurements are laborious and ignore the clinical presentation of the animal. The present study aimed to facilitate the assessment by establishing a body condition score. A total of 373 Hermann's Tortoises (Testudo hermanni) (n = 281 tortoises kept as pets in Germany and n = 92 tortoises originating from a free-ranging population (68) or a rearing station (24) in France) were examined and data (weight (g), carapace length (cm), width (cm), height (cm)) were recorded in a standard protocol between October 2020 and October 2021. A modified version of a body condition score for Mojave Desert Tortoises (Gopherus agassizii) (1 = cachectic, 3 = ideal, 5 = obese) was utilized and tested against pre-existing shell measurement systems (Jackson's ratio, body condition index, volume condition index, circumferential product). German captive tortoises were significantly heavier and larger than French specimens. In the Spearman's correlation matrix, the body condition score showed a statistically significant correlation with all measurement methods in the total population of captive tortoises (Testudo hermanni boettgeri), with a medium correlation strength, and a lack of correlation in free-ranging tortoises (Testudo hermanni hermanni). However, individual animal data suggested misleading results of mathematical equations in terms of body condition. Clinical evaluation of tortoises, including a body condition score, should be considered essential to provide good healthcare and should be an integral part of general examination.

Reduce, Replace, Refine: Determining A Posteriori Reference Intervals for Biochemistry in Hermann's Tortoise (Testudo hermanni).

Biochemical and hematological analyses are important for the assessment of animal health. However, for most wild species their use is hindered by the scarcity of reliable reference intervals. Indeed, collecting body fluids (e.g., blood, urine) in free-ranging animals is often technically challenging. Further, sampling many individuals would be essential to consider major sources of variations, such as species, populations, sex, age, and seasons. One alternative, according to the reduction, refinement, and replacement framework, is to establish reference intervals a posteriori using literature survey and unpublished data. We produced reference intervals for free-ranging Hermann's tortoises (Testudo hermanni), using analyses performed on blood samples collected in previous studies and conservation programs conducted in the field between 2010 and 2016 in southern France (n=195 individuals). Thirteen parameters were analyzed: packed-cell volume, blood concentrations of corticosterone, testosterone, glycemia, cholesterol, triglyceride, urea, uric acid, calcium, sodium, potassium, asparagine aminotransferase (AST), and alanine aminotransferases (ALT). Reference intervals for subgroups defined by sex and season were relevant for corticosterone, triglyceride, and calcium (sex) and cholesterol (season). Comparing our results with those obtained in captive individuals in Germany, except for urea and AST levels the intervals from both free-ranging versus captive tortoises were similar, suggesting that reference intervals established from captive individuals may be suitable for free-ranging populations in this species.

Proteomic insight into the venom composition of the largest European rear-fanged snake, Malpolon monspessulanus monspessulanus.

Snake envenomations constitute a worldwide neglected tropical disease, with the vast majority of lethal bites inflicted by front-fanged snakes from the viperid and elapid groups. Rear-fanged snakes (colubrids) were often considered harmless and as a result, are much less studied, but several documented deaths have suggested potent venom in this group as well. The largest European snake (Malpolon monspessulanus monspessulanus), known as the "Montpellier snake", is such a rear-fanged snake that belongs to the Lamprophiidae family. Its venom remains largely unknown but cases of envenomation with neurological symptoms have been reported. Here, we provide the first insights into the composition of its venom using mass spectrometry methods. First, liquid chromatography coupled mass spectrometry analysis of the manually collected venom samples reveals a complex profile, with the majority of masses encompassing the range 500-3000 Da, 4000-8000 Da, and 10 000-30 000 Da. Next, shotgun proteomics allowed the identification of a total of 42 different known families of proteins, including snake venom metalloproteinases, peptidase M1, and cysteine-rich secretory proteins, as the most prominent. Interestingly, three-finger toxins were not detected, suggesting that neurotoxicity may occur via other, yet to be determined, toxin types. Overall, our results provide the basis for a better understanding of the effects of a peculiar snake venom on human symptomatology, but also on the main prey consumed by this species.

Sex-specific ecophysiological responses to environmental fluctuations of free-ranging Hermann's tortoises: implication for conservation.

Physiological parameters provide indicators to evaluate how organisms respond to conservation actions. For example, individuals translocated during reinforcement programmes may not adapt to their novel host environment and may exhibit elevated chronic levels of stress hormones and/or decreasing body condition. Conversely, successful conservation actions should be associated with a lack of detrimental physiological perturbation. However, physiological references fluctuate over time and are influenced by various factors (e.g. sex, age, reproductive status). It is therefore necessary to determine the range of natural variations of the selected physiological metrics to establish useful baselines. This study focuses on endangered free-ranging Hermann's tortoises (Testudo hermanni hermanni), where conservation actions have been preconized to prevent extinction of French mainland populations. The influence of sex and of environmental factors (site, year and season) on eight physiological parameters (e.g. body condition, corticosterone concentrations) was assessed in 82 individuals from two populations living in different habitats. Daily displacements were monitored by radio-tracking. Most parameters varied between years and seasons and exhibited contrasting sex patterns but with no or limited effect of site. By combining behavioural and physiological traits, this study provides sex-specific seasonal baselines that can be used to monitor the health status of Hermann's tortoises facing environmental threats (e.g. habitat changes) or during conservation actions (e.g. translocation). These results might also assist in selection of the appropriate season for translocation.

Children prioritize virtual exotic biodiversity over local biodiversity.

Environmental education is essential to stem current dramatic biodiversity loss, and childhood is considered as the key period for developing awareness and positive attitudes toward nature. Children are strongly influenced by the media, notably the internet, about biodiversity and conservation issues. However, most media focus on a few iconic, appealing, and usually exotic species. In addition, virtual activities are replacing field experiences. This situation may curb children knowledge and concerns about local biodiversity. Focusing our analyses on local versus exotic species, we examined the level of knowledge and the level of diversity of the animals that French schoolchildren are willing to protect, and whether these perceptions are mainly guided by information available in the internet. For that, we collected and compared two complementary data sets: 1) a questionnaire was administered to schoolchildren to assess their knowledge and consideration to protect animals, 2) an internet content analysis (i.e. Google searching sessions using keywords) was performed to assess which animals are the most often represented. Our results suggest that the knowledge of children and their consideration to protect animal are mainly limited to internet contents, represented by a few exotic and charismatic species. The identification rate of local animals by schoolchildren was meager, suggesting a worrying disconnection from their local environment. Schoolchildren were more prone to protect "virtual" (unseen, exotic) rather than local animal species. Our results reinforce the message that environmental education must also focus on outdoor activities to develop conservation consciousness and concerns about local biodiversity.