Sylvain Ursenbacher

Site occupancy models in the analysis of environmental DNA presence/absence surveys : a case study of an emerging amphibian pathogen

Summary 1. The use of environmental DNA (eDNA) to detect species in aquatic environments such as ponds and streams is a powerful new technique with many benefits. However, species detection in eDNA-based surveys is likely to be imperfect, which can lead to underestimation of the distribution of a species. 2. Site occupancy models account for imperfect detection and can be used to estimate the proportion of sites where a species occurs from presence/absence survey data, making them ideal for the analysis of eDNA-based surveys. Imperfect detection can result from failure to detect the species during field work (e.g. by water samples) or during laboratory analysis (e.g. by PCR). 3. To demonstrate the utility of site occupancy models for eDNA surveys, we reanalysed a data set estimating the occurrence of the amphibian chytrid fungus Batrachochytrium dendrobatidis using eDNA. Our reanalysis showed that the previous estimation of species occurrence was low by 5–10%. Detection probability was best explained by an index of the number of hosts (frogs) in ponds. 4. Per-visit availability probability in water samples was estimated at 0� 45 (95% CRI 0� 32, 0� 58) and per-PCR detection probability at 0� 85 (95% CRI 0� 74, 0� 94), and six water samples from a pond were necessary for a cumulative detection probability >95%. A simulation study showed that when using site occupancy analysis, researchers need many fewer samples to reliably estimate presence and absence of species than without use of site occupancy modelling. 5. Our analyses demonstrate the benefits of site occupancy models as a simple and powerful tool to estimate detection and site occupancy (species prevalence) probabilities despite imperfect detection. As species detection from eDNA becomes more common, adoption of appropriate statistical methods, such as site occupancy models, will become crucial to ensure that reliable inferences are made from eDNA-based surveys.

The genus Atheris (Serpentes: Viperidae) in East Africa: phylogeny and the role of rifting and climate in shaping the current pattern of species diversity.

Past climatic and tectonic events are believed to have strongly influenced species diversity in the Eastern Afromontane Biodiversity Hotspot. We investigated the phylogenetic relationships and historical biogeography of the East African genus Atheris (Serpentes: Viperidae), and explored temporal and spatial relationships between Atheris species across Africa, and the impact of palaeoclimatic fluctuations and tectonic movements on cladogenesis of the genus. Using mitochondrial sequence data, the phylogeny of East African species of Atheris shows congruent temporal patterns that link diversification to major tectonic and aridification events within East Africa over the last 15million years (my). Our results are consistent with a scenario of a delayed direct west-east colonisation of the Eastern Arc Mountains of Atheris by the formation of the western rift. Based on the phylogenetic patterns, this terrestrial, forest-associated genus has dispersed into East Africa across a divided route, on both west-southeasterly and west-northeasterly directions (a C-shaped route). Cladogenesis in the Eastern Arc Mountains and Southern Highlands of Tanzania corresponds to late Miocene and Plio-Pleistocene climatic shifts. Taxonomically, our data confirmed the monophyly of Atheris as currently defined, and reveal four major East African clades, three of which occur in discrete mountain ranges. Possible cryptic taxa are identified in the Atheris rungweensis and A. ceratophora clades.

High Genetic Differentiation Among French Populations of the Orsini’s Viper (Vipera ursinii ursinii) Based on Mitochondrial and Microsatellite Data: Implications for Conservation Management

The Orsinis viper (Vipera ursinii) is one of the most threatened snakes in Europe due to its highly fragmented distribution and specific open environment (steppic habitat) requirement. French populations are isolated on top of mountain massifs of the southern Prealps/Alps. Mitochondrial sequences (cytochrome b) and 6 microsatellite loci have been used to estimate the levels of genetic diversity and isolation within and among 11 French fragmented populations (a total of 157 individuals). Eleven cytochrome b haplotypes with a limited divergence were observed (mean divergence between haplotypes: 0.31%). However, we detected considerable genetic differentiation among populations (global F(ST) = 0.76 and 0.26 for mitochondrial and nuclear DNA, respectively). Results indicate that 3 populations possibly went through a bottleneck and 1 population showed low genetic diversity compared with the others. Although a significant isolation by distance was detected for both markers, strong differentiation was also observed between geographically close populations, probably due to the ragged landscape that constitutes a serious barrier to gene flow owing to the limited dispersal capability of the viper. Despite some discrepancies between the 2 markers, 8 Management Units have been identified and should be considered for future management projects.

Male Reproductive Success and Multiple Paternity in Wild, Low-Density Populations of the Adder (Vipera berus)

We studied for the first time the occurrence of multiple paternity, male reproductive success, and neonate survival in wild, low-density adder (Vipera berus) populations using 13 microsatellite loci. Paternity was assigned for 15 clutches, collected during 3 years. Our data demonstrated that multiple paternity can occur at a high level (69%) in natural populations of V. berus, even if the density of adults is low. The high proportion of multiple sired clutches was comparable to the proportion observed in captive populations. Male reproductive success significantly increased with body length, and only the largest males successfully sired entire clutches. Finally, no relationship was detected between the number of fathers per clutch and neonate survival. These results suggest that multiple matings could be beneficial in populations with high level of inbreeding or low male fecundity.

Postglacial recolonization in a cold climate specialist in western Europe: patterns of genetic diversity in the adder (Vipera berus) support the central-marginal hypothesis

Understanding the impact of postglacial recolonization on genetic diversity is essential in explaining current patterns of genetic variation. The central–marginal hypothesis (CMH) predicts a reduction in genetic diversity from the core of the distribution to peripheral populations, as well as reduced connectivity between peripheral populations. While the CMH has received considerable empirical support, its broad applicability is still debated and alternative hypotheses predict different spatial patterns of genetic diversity. Using microsatellite markers, we analysed the genetic diversity of the adder (Vipera berus) in western Europe to reconstruct postglacial recolonization. Approximate Bayesian Computation (ABC) analyses suggested a postglacial recolonization from two routes: a western route from the Atlantic Coast up to Belgium and a central route from the Massif Central to the Alps. This cold‐adapted species likely used two isolated glacial refugia in southern France, in permafrost‐free areas during the last glacial maximum. Adder populations further from putative glacial refugia had lower genetic diversity and reduced connectivity; therefore, our results support the predictions of the CMH. Our study also illustrates the utility of highly variable nuclear markers, such as microsatellites, and ABC to test competing recolonization hypotheses.

Genetic differentiation in two European tree frog (Hyla arborea) metapopulations in contrasted landscapes of Western Switzerland

The survival of threatened species as the European tree frog (Hyla arborea) is strongly dependent on the genetic variability within populations, as well as gene flow between them. In Switzerland, only two sectors in its western part still harbour metapopulations. The first is characterised by a very heterogeneous and urbanized landscape, while the second is characterised by a uninterrupted array of suitable habitats. In this study, six microsatellite loci were used to establish levels of genetic differentiation among the populations from the two different locations. The results show that the metapopulations have: (i) weak levels of genetic differentiation (FST within metapopulation ≈ 0.04), (ii) no difference in levels of genetic structuring between them, (iii) significant (p = 0.019) differences in terms of genetic diversity (Hs) and observed heterozygozity (Ho), the metapopulation located in a disturbed landscape showing lower values. Our results suggest that even if the dispersal of H. arborea among contiguous ponds seems to be efficient in areas of heterogeneous landscape, a loss of genetic diversity can occur.

Influence of climate on the presence of colour polymorphism in two montane reptile species.

The coloration of ectotherms plays an important role in thermoregulation processes. Dark individuals should heat up faster and be able to reach a higher body temperature than light individuals and should therefore have benefits in cool areas. In central Europe, montane local populations of adder (Vipera berus) and asp viper (Vipera aspis) exhibit a varying proportion of melanistic individuals. We tested whether the presence of melanistic V. aspis and V. berus could be explained by climatic conditions. We measured the climatic niche position and breadth of monomorphic (including strictly patterned individuals) and polymorphic local populations, calculated their niche overlap and tested for niche equivalency and similarity. In accordance with expectations, niche overlap between polymorphic local populations of both species is high, and even higher than that of polymorphic versus monomorphic montane local populations of V. aspis, suggesting a predominant role of melanism in determining the niche of ectothermic vertebrates. However, unexpectedly, the niche of polymorphic local populations of both species is narrower than that of monomorphic ones, indicating that colour polymorphism does not always enable the exploitation of a greater variability of resources, at least at the intraspecific level. Overall, our results suggest that melanism might be present only when the thermoregulatory benefit is higher than the cost of predation.

Testing the Competitive Exclusion Principle using various niche parameters in a native (Natrix maura) and an introduced (N. tessellata) colubrid

Despite the increase of animal and plant introductions worldwide and the strong augmentation of the reptile trade, few invasive snake populations have been studied. Dice snakes (Natrix tessellata) were introduced to the shores of Lake Geneva (Switzerland) in the early 1920s, and are now well established. This region of introduction was previously inhabited by Viperine snakes (N. maura). Ever since these two species have been under monitoring (which began in 1996) the Viperine snake population has shown drastic decline. We examine here the possibility of trophic competition by analysing diet composition, prey size and trophic niche overlap. Spatial distribution is also assessed in order to address the question of spatial competitive exclusion. We found very similar diets, and thus a high trophic niche overlap, indicating no partitioning of the trophic resource. No arguments in favour of spatial competitive exclusion were found. Our study suggests that trophic competition may occur between the two natricines and that it may give an explanation for the drastic decline of the Viperine snake in this area. Other pathways potentially playing a role in the exclusion of the Viperine snake are discussed.

Systematics of the Swiss asp vipers : some implications for the European Vipera aspis (Linnaeus, 1758) complex (Serpentes Viperidae) : a tribute to Eugen Kramer

Following the opinion expressed by Kramerin 1971, it is commonly accepted that the V aspis complex is composed of 5 subspecies: V a. aspis (Linnaeus, 1758), V a. atra Meisner, 1820, V a. francisciredi Laurenti, 1768, V a. hugyi Schinz, 1834 and V a. zinnikeri Kramer, 1958. However, this point of view was recently challenged and a number of subspecies were elevated to species rank. We had the unique opportunity to study the sample employed by Kramer to revalidate the subspecies V a. atra and V a. francisciredi. Using Kramer`s determinations, but another set of characters, we reanalysed his sample, by applying the same statistical methods (discriminant and canonical analyses), in order to determine whether observed differences among taxa were sufficiently distinctive to warrant recognition as subspecies. The preliminary results of our morphological study allow us to admit the validity of V a. francisciredi Laurenti, 1768, but also to question the subspecific rank of V a. atra Meisner, 1820. These conclusions agree perfectly with the results of our previous genetic study.

Hybridisation between turtle subspecies: a case study with the European pond turtle (Emys orbicularis)

Genetic introgression has recently become an important concern for conservation genetics as it can occur between rare and related common species, between various genetic groups and between individuals from different origins. Our aims were: (i) to determine whether hybridisation or introgression occurs between Emys orbicularis subspecies in a natural contact zone in France; (ii) to determine the geographic extent of the contact zone between distinct subspecies in France; (iii) to evaluate at which level introgression occurs, and finally; (iv) to evaluate whether combining mitochondrial and nuclear genetic markers reflects the same contact zone as when only one genetic marker is used. Introgression was evaluated by genotyping biparentally inherited microsatellites and sequencing the maternally inherited cytochrome b gene of French populations. We demonstrated strong introgression between subspecies under natural conditions in the old contact zone in southern and eastern France. Our results corroborated that introgression reflects past natural events, but also demonstrated that human impact has altered these patterns. We finally confirmed that the combination of mitochondrial and nuclear genetic markers is more appropriate to reveal introgression than the use of only one genetic marker.