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Proteins encoded by antigen-processing genes (APGs) provide major histocompatibility complex (MHC) class I (MHC-I) with antigenic peptides. In mammals, polymorphic multigenic MHC-I family is served by monomorphic APGs, whereas in certain nonmammalian species both MHC-I and APGs are polymorphic and coevolve within stable haplotypes. Coevolution was suggested as an ancestral gnathostome feature, presumably enabling only a single highly expressed classical MHC-I gene. In this view coevolution, while optimizing some aspects of adaptive immunity, would also limit its flexibility by preventing the expansion of classical MHC-I into a multigene family. However, some nonmammalian taxa, such as salamanders, have multiple highly expressed MHC-I genes, suggesting either that coevolution is relaxed or that it does not prevent the establishment of multigene MHC-I. To distinguish between these two alternatives, we use salamanders (30 species from 16 genera representing six families) to test, within a comparative framework, a major prediction of the coevolution hypothesis: the positive correlation between MHC-I and APG diversity. We found that MHC-I diversity explained both within-individual and species-wide diversity of two APGs, TAP1 and TAP2, supporting their coevolution with MHC-I, whereas no consistent effect was detected for the other three APGs (PSMB8, PSMB9, and TAPBP). Our results imply that although coevolution occurs in salamanders, it does not preclude the expansion of the MHC-I gene family. Contrary to the previous suggestions, nonmammalian vertebrates thus may be able to accommodate diverse selection pressures with flexibility granted by rapid expansion or contraction of the MHC-I family, while retaining the benefits of coevolution between MHC-I and TAPs., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Many organisms live in networks of local populations connected by dispersing individuals, called spatially structured populations (SSPs), where the long-term persistence of the entire network is determined by the balance between 2 processes acting at the scale of local populations: extinction and colonization. When multiple threats act on an SSP, a comparison of the different factors determining local extinctions and colonizations is essential to plan sound conservation actions. We assessed the drivers of long-term population dynamics of multiple amphibian species at the regional scale. We used dynamic occupancy models within a Bayesian framework to identify the factors determining persistence and colonization of local populations. Because connectivity among patches is fundamental to SSPs dynamics, we considered 2 measures of connectivity acting on each focal patch: incidence of the focal species and incidence of invasive crayfish. We used meta-analysis to summarize the effect of different drivers at the community level. Persistence and colonization of local populations were jointly determined by factors acting at different scales. Persistence probability was positively related to the area and the permanence of wetlands, whereas it was negatively related to occurrence of fish. Colonization probability was highest in semipermanent wetlands and in sites with a high incidence of the focal species in nearby sites, whereas it showed a negative relationship with the incidence of invasive crayfish in the landscape. By analyzing long-term data on amphibian population dynamics, we found a strong effect of some classic features commonly used in SSP studies, such as patch area and focal species incidence. The presence of an invasive non-native species at the landscape scale emerged as one of the strongest drivers of colonization dynamics, suggesting that studies on SSPs should consider different connectivity measures more frequently, such as the incidence of predators, especially when dealing with biological invasions., (© 2020 Society for Conservation Biology.)

Long-term studies are essential to understand the impacts of global changes on the multiple facets of biological diversity. Here, we use distribution data for over 600 species of arthropods collected over 150 years from locations across Italy and test how multiple environmental stressors (climate, land use and human population density) influenced assemblage composition and functionality. By carefully reconstructing the temporal changes in these stressors, we explicitly tested how environmental changes can determine the observed changes in taxonomic and functional diversity. We found that rapid changes in precipitation destabilize the assemblages and maximize colonization and extinction rates, especially when coupled with changes in human population density (for taxonomy) or temperature (for functionality). Higher microclimatic heterogeneity increases the stability of biodiversity by reducing taxonomic and functional loss. Finally, changes in natural habitats increased colonization, influencing taxonomic nestedness and functional replacement. The integration of long-term datasets combining distributions, climate and traits may deepen our understanding of the processes underlying biodiversity responses to global-scale drivers., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)

Metabarcoding of bulk or environmental DNA has great potential for biomonitoring of freshwater environments. However, successful application of metabarcoding to biodiversity monitoring requires universal primers with high taxonomic coverage that amplify highly variable, short metabarcodes with high taxonomic resolution. Moreover, reliable and extensive reference databases are essential to match the outcome of metabarcoding analyses with available taxonomy and biomonitoring indices. Benthic invertebrates, particularly insects, are key taxa for freshwater bioassessment. Nevertheless, few studies have so far assessed markers for metabarcoding of freshwater macrobenthos. Here we combined in silico and laboratory analyses to test the performance of different markers amplifying regions in the 18S rDNA (Euka02), 16S rDNA (Inse01) and COI (BF1_BR2-COI) genes, and developed an extensive database of benthic macroinvertebrates of France and Europe, with a particular focus on key insect orders (Ephemeroptera, Plecoptera and Trichoptera). Analyses on 1,514 individuals representing different taxa of benthic macroinvertebrates showed very different amplification success across primer combinations. The Euka02 marker showed the highest universality, while the Inse01 marker showed excellent performance for the amplification of insects. BF1_BR2-COI showed the highest resolution, while the resolution of Euka02 was often limited. By combining our data with GenBank information, we developed a curated database including sequences representing 822 genera. The heterogeneous performance of the different primers highlights the complexity in identifying the best markers, and advocates for the integration of multiple metabarcodes for a more comprehensive and accurate understanding of ecological impacts on freshwater biodiversity., (© 2020 John Wiley & Sons Ltd.)

Macroinvertebrate assemblages are the most common bioindicators used for stream biomonitoring, yet the standard approach exhibits several time-consuming steps, including the sorting and identification of organisms based on morphological criteria. In this study, we examined if DNA metabarcoding could be used as an efficient molecular-based alternative to the morphology-based monitoring of streams using macroinvertebrates. We compared results achieved with the standard morphological identification of organisms sampled in 18 sites located on 15 French wadeable streams to results obtained with the DNA metabarcoding identification of sorted bulk material of the same macroinvertebrate samples, using read numbers (expressed as relative frequencies) as a proxy for abundances. In particular, we evaluated how combining and filtering metabarcoding data obtained from three different markers (COI: BF1-BR2, 18S: Euka02 and 16S: Inse01) could improve the efficiency of bioassessment. In total, 140 taxa were identified based on morphological criteria, and 127 were identified based on DNA metabarcoding using the three markers, with an overlap of 99 taxa. The threshold values used for sequence filtering based on the "best identity" criterion and the number of reads had an effect on the assessment efficiency of data obtained with each marker. Compared to single marker results, combining data from different markers allowed us to improve the match between biotic index values obtained with the bulk DNA versus morphology-based approaches. Both approaches assigned the same ecological quality class to a majority (86%) of the site sampling events, highlighting both the efficiency of metabarcoding as a biomonitoring tool but also the need for further research to improve this efficiency., (© 2020 John Wiley & Sons Ltd.)

The Aeolian wall lizard Podarcis raffonei is an island endemic that survives only on three tiny islets, and on the Capo Grosso peninsula of the Vulcano island, thus is among the European vertebrates with the smallest range and one of the most threatened by extinction. This species is declining due to competition and hybridization with the non-native lizard Podarcis siculus, but a regular monitoring program is lacking. Here we assessed the size and status of the Capo Grosso population of P. raffonei on Vulcano. In September 2015 we captured 30 individuals showing the typical brown phenotype of P. raffonei, while one single male showed a green phenotype, apparently intermediate between P. raffonei and the non-native Podarcis siculus. In May 2017, only 47% of 131 individuals showed the typical brown phenotype (P. raffonei-like) and 53% showed the green phenotype (P. siculus-like). Based on N-mixture models and removal sampling the estimated size of the Capo Grosso population was of 800-1300 individuals in 2017, being similar to 2015; available data suggest that the total range of the species could be as small as 2 ha. The frequency of individuals with the typical P. raffonei phenotype dramatically dropped between two samplings with a parallel increase of individuals displaying the green phenotype. Observation on outdoor captive-bred individuals demonstrates plasticity for colouration in P. raffonei individuals from Capo Grosso, with several individuals showing the typical brown pattern in September 2017 and a green pattern in March 2021. Non-exclusive hypotheses, including hybridization with P. siculus and plasticity in colour pattern of P. raffonei, are discussed to explain the phenotypic shifts of the P. raffonei population of Capo Grosso. While genomic evidence is required to reach conclusions and investigate eventual hybridization, it is urgent to undertake a programme for the monitoring and management of this lizard., Competing Interests: The authors have declared that no competing interests exist.

European plethodontid salamanders (genus Speleomantes; formerly Hydromantes) are a group of eight strictly protected amphibian species which are sensitive to human-induced environmental changes. Long-term monitoring is highly recommended to evaluate their status and to assess potential threats. Here we used two low-impact methodologies to build up a large dataset on two mainland Speleomantes species (S. strinatii and S. ambrosii), which represents an update to two previously published datasets, but also includes several new populations. Specifically, we provide a set of 851 high quality images and a table gathering stomach contents recognized from 560 salamanders. This dataset offers the opportunity to analyse phenotypic traits and stomach contents of eight populations belonging to two Speleomantes species. Furthermore, the data collection performed over different periods allows to expand the potential analyses through a wide temporal scale, allowing long-term studies.

Direct evidence of ancient human occupation is typically established through archaeological excavation. Excavations are costly and destructive, and practically impossible in some lake and wetland environments. We present here an alternative approach, providing direct evidence from lake sediments using DNA metabarcoding, steroid lipid biomarkers (bile acids) and from traditional environmental analyses. Applied to an early Medieval Celtic settlement in Ireland (a crannog) this approach provides a site chronology and direct evidence of human occupation, crops, animal farming and on-site slaughtering. This is the first independently-dated, continuous molecular archive of human activity from an archeological site, demonstrating a link between animal husbandry, food resources, island use. These sites are under threat but are impossible to preserve in-situ so this approach can be used, with or without excavation, to produce a robust and full site chronology and provide direct evidence of occupation, the use of plants and animals, and activities such as butchery.

The species-area relationship (SAR) is one of the most well-established scaling patterns in ecology. Its implications for understanding how communities change across spatial gradients are numerous, including the effects of habitat loss on biodiversity. However, ecological communities are not mere collections of species. They are the result of interactions between these species forming complex networks that tie them together. Should we aim to grasp the spatial scaling of biodiversity as a whole, it is fundamental to understand the changes in the structure of interaction networks with area. In spite of a few empirical and theoretical studies that address this challenge, we still do not know much about how network structure changes with area, or what are the main environmental drivers of these changes. Here, using the meta-network of potential interactions between all terrestrial vertebrates in Europe (1140 species and 67 201 feeding interactions), we analysed network-area relationships (NARs) that summarize how network properties scale with area. We do this across ten biogeographical regions, which differ in environmental characteristics. We found that the spatial scaling of network complexity strongly varied across biogeographical regions. However, once the variation in SARs was accounted for, differences in the shape of NARs vanished. On the other hand, the proportion of species across trophic levels remained remarkably constant across biogeographical regions and spatial scales, despite the great variation in species richness. Spatial variation in mean annual temperature and habitat clustering were the main environmental determinants of the shape of both SARs and NARs across Europe. Our results suggest new avenues in the exploration of the effects of environmental factors on the spatial scaling of biodiversity. We argue that NARs can provide new insights to analyse and understand ecological communities., Competing Interests: Conflicts of interest – None of the authors have any conflict of interest.

Since the last glacial maximum, soil formation related to ice-cover shrinkage has been one major sink of carbon accumulating as soil organic matter (SOM), a phenomenon accelerated by the ongoing global warming. In recently deglacierized forelands, processes of SOM accumulation, including those that control carbon and nitrogen sequestration rates and biogeochemical stability of newly sequestered carbon, remain poorly understood. Here, we investigate the build-up of SOM during the initial stages (up to 410 years) of topsoil development in 10 glacier forelands distributed on four continents. We test whether the net accumulation of SOM on glacier forelands (i) depends on the time since deglacierization and local climatic conditions (temperature and precipitation); (ii) is accompanied by a decrease in its stability and (iii) is mostly due to an increasing contribution of organic matter from plant origin. We measured total SOM concentration (carbon, nitrogen), its relative hydrogen/oxygen enrichment, stable isotopic ( 13 C, 15 N) and carbon functional groups (C-H, C=O, C=C) compositions, and its distribution in carbon pools of different thermal stability. We show that SOM content increases with time and is faster on forelands experiencing warmer climates. The build-up of SOM pools shows consistent trends across the studied soil chronosequences. During the first decades of soil development, the low amount of SOM is dominated by a thermally stable carbon pool with a small and highly thermolabile pool. The stability of SOM decreases with soil age at all sites, indicating that SOM storage is dominated by the accumulation of labile SOM during the first centuries of soil development, and suggesting plant carbon inputs to soil (SOM depleted in nitrogen, enriched in hydrogen and in aromatic carbon). Our findings highlight the potential vulnerability of SOM stocks from proglacial areas to decomposition and suggest that their durability largely depends on the relative contribution of carbon inputs from plants., (© 2020 The Authors. Global Change Biology published by John Wiley & Sons Ltd.)

Environmental factors are well known to affect spatio-temporal patterns of infectious disease outbreaks, but whether the rapid spread of COVID-19 across the globe is related to local environmental conditions is highly debated. We assessed the impact of environmental factors (temperature, humidity and air pollution) on the global patterns of COVID-19 early outbreak dynamics during January-May 2020, controlling for several key socio-economic factors and airport connections. We showed that during the earliest phase of the global outbreak (January-March), COVID-19 growth rates were non-linearly related to climate, with fastest spread in regions with a mean temperature of ca. 5 °C, and in the most polluted regions. However, environmental effects faded almost completely when considering later outbreaks, in keeping with the progressive enforcement of containment actions. Accordingly, COVID-19 growth rates consistently decreased with stringent containment actions during both early and late outbreaks. Our findings indicate that environmental drivers may have played a role in explaining the early variation among regions in disease spread. With limited policy interventions, seasonal patterns of disease spread might emerge, with temperate regions of both hemispheres being most at risk of severe outbreaks during colder months. Nevertheless, containment measures play a much stronger role and overwhelm impacts of environmental variation, highlighting the key role for policy interventions in curbing COVID-19 diffusion within a given region. If the disease will become seasonal in the next years, information on environmental drivers of COVID-19 can be integrated with epidemiological models to inform forecasting of future outbreak risks and improve management plans., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

High-throughput sequencing (HTS) is increasingly being used for the characterization and monitoring of biodiversity. If applied in a structured way, across broad geographical scales, it offers the potential for a much deeper understanding of global biodiversity through the integration of massive quantities of molecular inventory data generated independently at local, regional and global scales. The universality, reliability and efficiency of HTS data can potentially facilitate the seamless linking of data among species assemblages from different sites, at different hierarchical levels of diversity, for any taxonomic group and regardless of prior taxonomic knowledge. However, collective international efforts are required to optimally exploit the potential of site-based HTS data for global integration and synthesis, efforts that at present are limited to the microbial domain. To contribute to the development of an analogous strategy for the nonmicrobial terrestrial domain, an international symposium entitled "Next Generation Biodiversity Monitoring" was held in November 2019 in Nicosia (Cyprus). The symposium brought together evolutionary geneticists, ecologists and biodiversity scientists involved in diverse regional and global initiatives using HTS as a core tool for biodiversity assessment. In this review, we summarize the consensus that emerged from the 3-day symposium. We converged on the opinion that an effective terrestrial Genomic Observatories network for global biodiversity integration and synthesis should be spatially led and strategically united under the umbrella of the metabarcoding approach. Subsequently, we outline an HTS-based strategy to collectively build an integrative framework for site-based biodiversity data generation., (© 2021 The Authors. Molecular Ecology published by John Wiley & Sons Ltd.)

Tris(1-chloro-2-propyl)phosphate (TCPP) is the most common chlorinated organophosphorus flame retardant in seawater. Due to its chemical features and abundance, TCPP has been classified as a high hazard, and restrictions of use have been set in multiple countries. Despite TCPP being highly present in the marine environment, only a few studies have explored the TCPP impact on the development of marine invertebrates. Ascidians are important invertebrate members of benthic marine communities and reliable model systems for ecotoxicological research. The aim of this study was to assess the adverse effects of TCPP exposure on the embryogenesis of the ascidian Ciona intestinalis. Our results showed that this pollutant affected both muscles and nervous system development. Malformations appeared similar to those reported in other animal models for other flame retardants, suggesting that these molecules could share a common mechanism of action and induce a mixture effect when simultaneously present in the aquatic environment even at sub-teratogenic concentrations., (© 2021 Wiley Periodicals LLC.)

Proteins encoded by antigen-processing genes (APGs) prepare antigens for presentation by the major histocompatibility complex class I (MHC I) molecules. Coevolution between APGs and MHC I genes has been proposed as the ancestral gnathostome condition. The hypothesis predicts a single highly expressed MHC I gene and tight linkage between APGs and MHC I. In addition, APGs should evolve under positive selection, a consequence of the adaptive evolution in MHC I. The presence of multiple highly expressed MHC I genes in some teleosts, birds, and urodeles appears incompatible with the coevolution hypothesis. Here, we use urodele amphibians to test two key expectations derived from the coevolution hypothesis: 1) the linkage between APGs and MHC I was studied in Lissotriton newts and 2) the evidence for adaptive evolution in APGs was assessed using 42 urodele species comprising 21 genera from seven families. We demonstrated that five APGs (PSMB8, PSMB9, TAP1, TAP2, and TAPBP) are tightly linked (<0.5 cM) to MHC I. Although all APGs showed some codons under episodic positive selection, we did not find a pervasive signal of positive selection expected under the coevolution hypothesis. Gene duplications, putative gene losses, and divergent allelic lineages detected in some APGs demonstrate considerable evolutionary dynamics of APGs in salamanders. Overall, our results indicate that if coevolution between APGs and MHC I occurred in urodeles, it would be more complex than envisaged in the original formulation of the hypothesis., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

The worldwide biodiversity crisis with the resulting need to increase species protection has led researchers to pursue and select survey methods that guarantee the best quality of data and produce the least negative effects on wild animals. Plethodontids are the most diverse family of salamanders; all species are very sensitive to human handling and noninvasive, but accurate, measurement methods are needed to reduce researchers' impact. Here, we tested the reliability of a noninvasive post hoc method in estimating the snout-vent length (SVL) from photographs showing salamanders' dorsal view. The correlation between the estimated snout-vent length (SVL e ) and the conventional SVL was high ( R 2 m  = .81), and no significant difference occurred between operators with different experience. Finally, we list the numerous advantages for the use of SVL e in terms of data quality and in reducing the stress caused to wild animals., Competing Interests: None declared., (© 2020 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.)

The COVID-19 pandemic zoonosis has determined extensive lockdowns worldwide that provide an unprecedented opportunity to understand how large-scale shifts of human activities can impact wildlife. We addressed the impacts of the COVID-19 lockdown on wildlife in Italy, the first European country that performed a countrywide lockdown, and identified potentially beneficial and negative consequences for wildlife conservation and management. We combined a qualitative analysis of social media information with field data from multiple taxa, data from citizen science projects, and questionnaires addressed to managers of protected areas. Both social media information and field data suggest that a reduction of human disturbance allowed wildlife to exploit new habitats and increase daily activity. The field data confirmed some positive effects on wildlife conservation, such as an increase in species richness in temporarily less-disturbed habitats, a higher breeding success of an aerial insectivorous bird, and reduction of road-killing of both amphibians and reptiles. Despite some positive effects, our data also highlighted several negative impacts of the COVID-19 crisis on wildlife. The lower human disturbance linked to lockdown was in fact beneficial for invasive alien species. Results from questionnaires addressed to managers of protected areas highlighted that the COVID-19 lockdown interrupted actions for the control of invasive alien species, and hampered conservation activities targeting threatened taxa. Furthermore, the reduction of enforcement could cause a surge of illegal killing of wildlife. The COVID-19 crisis, besides having deep socio-economic impacts, might profoundly affect wildlife conservation, with potentially long-lasting effects., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2020 Elsevier Ltd. All rights reserved.)

Individual diet specialization (IS) has important community- and population-level implications and its ecological drivers are actively investigated. Here, to test the hypothesis that local environmental conditions may influence IS in wild populations, we analyzed the stomach contents of 395 individuals from eight populations of five allopatric species of European cave salamanders (genus Hydromantes). We assessed whether their degree of individual diet specialization (1) scaled positively with the respective niche widths, in agreement with Van Valen's niche variation hypothesis (NVH), and (2) could be predicted by satellite-derived climatic and vegetation characteristics of the sites where the populations live. Consistent with the NVH, the degree of individual diet specialization increased with the populations' total niche width. Furthermore, two variables describing local nonarboreal vegetation cover and habitat heterogeneity successfully predicted the variation in individual specialization across the eight populations. Climatic factors had a generally low predictive power, with individual specialization in low- and high-elevation populations showing contrasting patterns of co-variation with air temperature in the warmest quarter of the year. However, independently from elevation, specialization peaked under conditions of high nonarboreal vegetation cover and high precipitation regimes. We discussed the results against two mutually nonexclusive scenarios hypothesizing different mechanisms linking environmental factors to salamanders' trophic strategy at an individual and population level. We concluded that satellite-derived climatic and vegetation variables to date generally adopted to model Grinnellian niches might also be useful in predicting spatial variations in dietary habits of populations, that is, their Eltonian niches., (© 2020 by the Ecological Society of America.)

Invasive predators can exert strong selection on native populations. If selection is strong enough, populations could lose the phenotypic variation caused by adaptation to heterogeneous environments. We compare frog tadpoles prior to and 14 years following invasion by crayfish. Prior to the invasion, populations differed in their intrinsic developmental rate, with tadpoles from cold areas reaching metamorphosis sooner than those from warm areas. Following the invasion, tadpoles from invaded populations develop faster than those from non-invaded populations. This ontogenetic shift overwhelmed the intraspecific variation between populations in a few generations, to the point where invaded populations develop at a similar rate regardless of climate. Rapid development can have costs, as fast-developing froglets have a smaller body size and poorer jumping performance, but compensatory growth counteracts some costs of development acceleration. Strong selection by invasive species can disrupt local adaptations by dampening intraspecific phenotypic variation, with complex consequences on lifetime fitness.

eDNA refers to DNA extracted from an environmental sample with the goal of identifying the occurrence of past or current biological communities in aquatic and terrestrial environments. However, there is currently a lack of knowledge regarding the soil memory effect and its potential impact on lake sediment eDNA records. To investigate this issue, two contrasted sites located in cultivated environments in France were studied. In the first site, soil samples were collected (n = 30) in plots for which the crop rotation history was documented since 1975. In the second site, samples were collected (n = 40) to compare the abundance of currently observed taxa versus detected taxa in cropland and other land uses. The results showed that the last cultivated crop was detected in 100% of the samples as the most abundant. In addition, weeds were the most abundant taxa identified in both sites. Overall, these results illustrate the potential of eDNA analyses for identifying the recent (< 10 years) land cover history of soils and outline the detection of different taxa in cultivated plots. The capacity of detection of plant species grown on soils delivering sediments to lacustrine systems is promising to improve our understanding of sediment transfer processes over short timescales.

European Hydromantes are a group of eight salamander species often occurring in subterranean habitats, which are a difficult environment to explore. All Hydromantes are strictly protected species and thus, low-impact methodologies to study these salamanders are strongly needed. Here we used a photographic technique to produce a large dataset of European Hydromantes, providing standardised pictures of 1,052 individuals belonging to the eight species, including hybrids as well. With our methodology we were able to reduce the handling time of individuals, and produce high quality pictures useful to investigate multiple life traits of these endangered species. Furthermore, the standardised photos provided here can be used for future comparisons of individuals from the surveyed populations.

Microhabitat selection is a key component of amphibian breeding biology and can be modulated in response to the features of breeding sites and the presence of predators. Despite invasive alien species being among the major threats to amphibians, there is limited information on the role of invasive species in shaping amphibians' breeding microhabitat choice. The invasive red swamp crayfish ( Procambarus clarkii) is a major predator of amphibians' larvae, including those of the brown frogs Rana dalmatina and Rana latastei . Although qualitative information about the spawning site preferences and breeding microhabitat choice of brown frogs is available in the literature, only a few studies performed quantitative analyses, and the relationship between microhabitat choice and the presence of alien predators has not been investigated yet. The aims of this study were: (1) to characterize the microhabitats selected for clutch deposition by R. dalmatina and R. latastei and (2) to test if the position and the aggregation of egg clutches differ in sites invaded or not invaded by P. clarkii . During spring 2017, we surveyed multiple times 15 breeding sites of both brown frogs in Northern Italy; in each site we assessed the features of the microhabitat where each egg clutch was laid, considering its position (distance from the shore, depth of the water column) and the degree of aggregation of clutches. In each site we also assessed the presence/absence of the invasive crayfish and the relative abundance in the breeding period. We detected egg clutches in all sites; the crayfish occurred in eight ponds. Our results showed substantial differences between the spawning microhabitat features of the two brown frogs: Rana latastei clutches showed a higher degree of aggregation and were associated with deeper areas of the ponds , while Rana dalmatina deposited more spaced out clutches in areas of the ponds that were less deep. For both species, spawning microhabitat features were not significantly different between sites with and without P. clarkii . Although we did not detect behavioural responses to P. clarkii in the choice of spawning microhabitat , additional studies are required to assess whether these frogs modulate other behavioural traits (e.g. during larval development) in response to the invasive predator., Competing Interests: The authors declare there are no competing interests., (©2020 Romagnoli et al.)

Environmental change is rapidly accelerating, and many species will need to adapt to survive 1 . Ensuring that protected areas cover populations across a broad range of environmental conditions could safeguard the processes that lead to such adaptations 1-3 . However, international conservation policies have largely neglected these considerations when setting targets for the expansion of protected areas 4 . Here we show that-of 19,937 vertebrate species globally 5-8 -the representation of environmental conditions across their habitats in protected areas (hereafter, niche representation) is inadequate for 4,836 (93.1%) amphibian, 8,653 (89.5%) bird and 4,608 (90.9%) terrestrial mammal species. Expanding existing protected areas to cover these gaps would encompass 33.8% of the total land surface-exceeding the current target of 17% that has been adopted by governments. Priority locations for expanding the system of protected areas to improve niche representation occur in global biodiversity hotspots 9 , including Colombia, Papua New Guinea, South Africa and southwest China, as well as across most of the major land masses of the Earth. Conversely, we also show that planning for the expansion of protected areas without explicitly considering environmental conditions would marginally reduce the land area required to 30.7%, but that this would lead to inadequate niche representation for 7,798 (39.1%) species. As the governments of the world prepare to renegotiate global conservation targets, policymakers have the opportunity to help to maintain the adaptive potential of species by considering niche representation within protected areas 1,2 .

Background: Systematic data collection on species and their exploited environments is of key importance for conservation studies. Within the less-known environments, the subterranean ones are neither easy to be studied, nor to be explored. Subterranean environments house a wide number of specialised organisms, many of which show high sensitivity to habitat alteration. Despite the undeniable importance to monitor the status of the subterranean biodiversity, standardised methodologies to record biotic and abiotic data in these environments are still not fully adopted, impeding therefore the creation of comparable datasets useful for monitoring the ecological condition in the subterranean environments and for conservation assessment of related species., New Information: In this work we describe a methodology allowing the collection of standardised abiotic and biotic data in subterranean environments. To show this, we created a large dataset including information on environmental features (morphology and microclimate) and abundance of five predators (one salamander, three spiders and one snail) occurring in seven caves of the Monte Albo (Sardinia, Italy), an important biodiversity hotspot. We performed 77 surveys on 5,748 m 2 of subterranean environments througout a year, recording 1,695 observations of the five cave predators. The fine-scale data collection adopted in our methodology allowed us to record detailed information related to both morphology and microclimate of the cave inner environment. Furthermore, this method allows us to account for species-imperfect detection when recording presence/abundance data., (Enrico Lunghi, Claudia Corti, Manuela Mulargia, Yahui Zhao, Raoul Manenti, Gentile Francesco Ficetola, Michael Veith.)

Over the last decade, an increasing number of studies have used lake sediment DNA to trace past landscape changes, agricultural activities or human presence. However, the processes responsible for lake sediment formation and sediment properties might affect DNA records via taphonomic and analytical processes. It is crucial to understand these processes to ensure reliable interpretations for "palaeo" studies. Here, we combined plant and mammal DNA metabarcoding analyses with sedimentological and geochemical analyses from three lake-catchment systems that are characterised by different erosion dynamics. The new insights derived from this approach elucidate and assess issues relating to DNA sources and transfer processes. The sources of eroded materials strongly affect the "catchment-DNA" concentration in the sediments. For instance, erosion of upper organic and organo-mineral soil horizons provides a higher amount of plant DNA in lake sediments than deep horizons, bare soils or glacial flours. Moreover, high erosion rates, along with a well-developed hydrographic network, are proposed as factors positively affecting the representation of the catchment flora. The development of open and agricultural landscapes, which favour the erosion, could thus bias the reconstructed landscape trajectory but help the record of these human activities. Regarding domestic animals, pastoral practices and animal behaviour might affect their DNA record because they control the type of source of DNA ("point" vs. "diffuse").

Understanding and counteracting biodiversity losses requires quantitative knowledge on species distribution and abundance across space and time, as well as integrated and interoperable information on climate conditions and climatic changes. In this paper we developed a new biodiversity-climate database for Italy, ClimCKmap, based on the critical analysis, quality estimation and subsequent integration of the CKmap database with several high-resolution climate datasets. The original database was quality-checked for errors in toponym, species name and dating; the retained records were georeferenced and their distribution polygonised via Voronoi tessellation. We then integrated the species distribution information with several high-resolution climatic datasets: average monthly minimum and maximum temperature and total monthly precipitation were reconstructed for each Voronoi cell and year. The resulting database contains 268,977 occurrence records from 8,445 binomials and 16,332 localities, dating between 1680 and 2006 CE. This dataset, fully available at https://doi.org/10.6084/m9.figshare.7906739.v4 and http://hdl.handle.net/21.11125/a91f85cb-befd-4e14-8e83-24f17c4a0491 , represents the largest, fully quality-checked, spatially, temporally and climatically explicit distribution database ever assembled for the Italian fauna, now ready for scientific exploitation.

The continuous decline of biodiversity is determined by the complex and joint effects of multiple environmental drivers. Still, a large part of past global change studies reporting and explaining biodiversity trends have focused on a single driver. Therefore, we are often unable to attribute biodiversity changes to different drivers, since a multivariable design is required to disentangle joint effects and interactions. In this work, we used a meta-regression within a Bayesian framework to analyze 843 time series of population abundance from 17 European amphibian and reptile species over the last 45 years. We investigated the relative effects of climate change, alien species, habitat availability, and habitat change in driving trends of population abundance over time, and evaluated how the importance of these factors differs across species. A large number of populations (54%) declined, but differences between species were strong, with some species showing positive trends. Populations declined more often in areas with a high number of alien species, and in areas where climate change has caused loss of suitability. Habitat features showed small variation over the last 25 years, with an average loss of suitable habitat of 0.1%/year per population. Still, a strong interaction between habitat availability and the richness of alien species indicated that the negative impact of alien species was particularly strong for populations living in landscapes with less suitable habitat. Furthermore, when excluding the two commonest species, habitat loss was the main correlate of negative population trends for the remaining species. By analyzing trends for multiple species across a broad spatial scale, we identify alien species, climate change, and habitat changes as the major drivers of European amphibian and reptile decline., (© 2019 John Wiley & Sons Ltd.)

An excessive supply of sediment is observed in numerous rivers across the world where it leads to deleterious impacts. Information on the sources delivering this material to waterbodies is required to design effective management measures, and sediment tracing or fingerprinting techniques are increasingly used to quantify the amount of sediment derived from different sources. However, the current methods used to identify the land use contributions to sediment have a limited discrimination power. Here, we investigated the potential of environmental DNA (eDNA) to provide more detailed information on the plant species found in sediment source areas as a next generation fingerprint. To this end, flood sediment deposits (n = 12) were collected in 2017 in two catchments impacted by the Fukushima radioactive fallout along differing river sections draining forests, cropland or a mix of both land uses. Conventional fingerprints (i.e. fallout radionuclides and organic matter properties) were also measured in these samples. The conventional fingerprint model results showed that most sediment samples contained a dominant proportion of subsoil material. Nevertheless, the eDNA information effectively discriminated the three above-mentioned groups of sediment, with the dominance of tree, shrub and fern species in sediment sampled in rivers draining forests versus a majority of grass, algae and cultivated plant species in sediment collected in rivers draining cropland. Based on these encouraging results, future research should examine the potential of eDNA in mixed land use catchments where the contribution of topsoil to sediment dominates and where the cultivation of land has not been abandoned in order to better characterize the memory effect of eDNA in soils and sediment., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Site occupancy-detection models (SODMs) are statistical models widely used for biodiversity surveys where imperfect detection of species occurs. For instance, SODMs are increasingly used to analyse environmental DNA (eDNA) data, taking into account the occurrence of both false-positive and false-negative errors. However, species occurrence data are often characterized by spatial and temporal autocorrelation, which might challenge the use of standard SODMs. Here we reviewed the literature of eDNA biodiversity surveys and found that most of studies do not take into account spatial or temporal autocorrelation. We then demonstrated how the analysis of data with spatial or temporal autocorrelation can be improved by using a conditionally autoregressive SODM, and show its application to environmental DNA data. We tested the autoregressive model on both simulated and real data sets, including chronosequences with different degrees of autocorrelation, and a spatial data set on a virtual landscape. Analyses of simulated data showed that autoregressive SODMs perform better than traditional SODMs in the estimation of key parameters such as true-/false-positive rates and show a better discrimination capacity (e.g., higher true skill statistics). The usefulness of autoregressive SODMs was particularly high in data sets with strong autocorrelation. When applied to real eDNA data sets (eDNA from lake sediment cores and freshwater), autoregressive SODM provided more precise estimation of true-/false-positive rates, resulting in more reasonable inference of occupancy states. Our results suggest that analyses of occurrence data, such as many applications of eDNA, can be largely improved by applying conditionally autoregressive specifications to SODMs., (© 2018 John Wiley & Sons Ltd.)

The evolutionary and ecological importance of neutral and adaptive genetic diversity is widely recognized. Nevertheless, genetic diversity is rarely assessed for conservation planning, which often implicitly assumes a positive correlation between species and genetic diversity. Multiple drivers can cause the co-variation between the genetic diversity of one species and the richness of the whole communities, and explicit tests are needed to identify the processes that can determine species-genetic diversity correlations (SGDCs). Here, we tested whether intrapopulation genetic diversity (at neutral loci) and species richness co-vary in the amphibian communities of a southern Alpine region (Trentino, Italy), using the common frog (Rana temporaria) as focal species for the study of genetic diversity. We also analysed ecological similarity, niche overlap and interspecific interactions between the species, to unravel the processes determining SGDC. The neutral genetic diversity of common frogs was negatively related to species richness. The negative SGDC was probably due to an opposite influence of environmental gradients on the two levels of biodiversity, since the focal species and the other amphibians differ in ecological preferences, particularly in terms of thermal optimum. Conversely, we did not find evidence for a role of interspecific interactions in the negative SGDC. Our findings stress that species richness cannot be used as a universal proxy for genetic diversity, and only combining SGDC with analyses on the determinants of biodiversity can allow to identify the processes determining the relationships between genetic and species diversity., (© 2018 John Wiley & Sons Ltd.)

Nano-encapsulation is a technology used to pack substances in order to enhance their stability and bioavailability, but this packing may interact with living systems, causing unexpected toxicity. Vitamin A (vit A) is a substance that has received attention, because in developed countries, the increasing availability of supplements is leading to its excessive intake. This study aims to compare teratogenic effects caused by exposure to the traditional formulation of vit A versus nano-encapsulated vit A. We used ascidian embryos as an alternative model. Ascidians are marine organisms closely related to vertebrates that share with them a body plan and developmental programme, including the morphogenetic role of retinoic acid (RA). Our data showed that the adverse effects of exposure to the same concentration of the two formulations were different, suggesting that the nano-encapsulation increased the bioavailability of the molecule, which could be better absorbed and metabolised to RA, the effective teratogenic substance.

The trophic niche is a life trait that identifies the consumer's position in a local food web. Several factors, such as ontogeny, competitive ability and resource availability contribute in shaping species trophic niches. To date, information on the diet of European Hydromantes salamanders are only available for a limited number of species, no dietary studies have involved more than one species of the genus at a time, and there are limited evidences on how multiple factors interact in determining diet variation. In this study we examined the diet of multiple populations of six out of the eight European cave salamanders, providing the first data on the diet for five of them. In addition, we assessed whether these closely related generalist species show similar diet and, for each species, we tested whether season, age class or sex influence the number and the type of prey consumed. Stomach condition (empty/full) and the number of prey consumed were strongly related to seasonality and to the activity level of individuals. Empty stomachs were more frequent in autumn, in individuals far from cave entrance and in juveniles. Diet composition was significantly different among species. Hydromantes imperialis and H. supramontis were the most generalist species; H. flavus and H. sarrabusensis fed mostly on Hymenoptera and Coleoptera Staphylinidae, while H. genei and H. ambrosii mostly consumed Arachnida and Endopterygota larvae. Furthermore, we detected seasonal shifts of diet in the majority of the species examined. Conversely, within each species, we did not find diet differences between females, males and juveniles. Although being assumed to have very similar dietary habits, here Hydromantes species were shown to be characterized by a high divergence in diet composition and in the stomach condition of individuals., Competing Interests: The authors have declared that no competing interests exist.

Life history changes may change resource use. Such shifts are not well understood in the dung beetles, despite recognised differences in larval and adult feeding ability. We use the flightless dung beetle Circellium bacchus to explore such shifts, identifying dung sources of adults using DNA metabarcoding, and comparing these with published accounts of larval dung sources. C. bacchus is traditionally considered to specialise on the dung of large herbivores for both larval and adult feeding. We successfully extracted mammal DNA from 151 adult C. bacchus fecal samples, representing 16 mammal species (ranging from elephants to small rodents), many of which are hitherto undescribed in the diet. Adult C. bacchus showed clear dung source preferences, especially for large herbivores inhabiting dense-cover vegetation. Our approach also confirmed the presence of cryptic taxa in the study area, and we propose that this may be used for biodiversity survey and monitoring purposes. Murid rodent feces were the most commonly fed-upon dung source (77.5%) for adult C. bacchus, differing markedly from the large and megaherbivore dung sources used for larval rearing. These findings support the hypothesis of life history-specific shifts in resource use in dung beetles, and reveal a hitherto unsuspected, but ecologically important, role of these dung beetles in consuming rodent feces. The differences in feeding abilities of the larval and adult life history stages have profound consequences for their resource use and foraging strategies, and hence the ecological role of dung beetles. This principle and its ecological consequences should be explored in other scarabaeids.

The extent to which closely related species share similar niches remains highly debated. Ecological niches are increasingly analysed by combining distribution records with broad-scale climatic variables, but interactions between species and their environment often occur at fine scales. The idea that macroscale analyses correctly represent fine-scale processes relies on the assumption that average climatic variables are meaningful predictors of processes determining species persistence, but tests of this hypothesis are scarce. We compared broad- and fine-scale (microhabitat) approaches by analyzing the niches of European plethodontid salamanders. Both the microhabitat and the macroecological approaches identified niche differences among species, but the correspondence between micro- and macroecological niches was weak. When exploring niche evolution, the macroecological approach suggested a close relationship between niche and phylogenetic history, but this relationship did not emerge in fine-scale analyses. The apparent pattern of niche evolution emerging in broad-scale analyses likely was the by-product of related species having closely adjacent ranges. The environment actually experienced by most of animals is more heterogeneous than what is apparent from macro-scale predictors, and a better combination between macroecological and fine-grained data may be a key to obtain robust ecological generalizations.

Accurate measures of species abundance are essential to identify conservation strategies. N-mixture models are increasingly used to estimate abundance on the basis of species counts. In this study we tested whether abundance estimates obtained using N-mixture models provide consistent results with more traditional approaches requiring capture (capture-mark recapture and removal sampling). We focused on endemic, threatened species of amphibians and reptiles in Italy, for which accurate abundance data are needed for conservation assessments: the Lanza's Alpine salamander Salamandra lanzai, the Ambrosi's cave salamander Hydromantes ambrosii and the Aeolian wall lizard Podarcis raffonei. In visual counts, detection probability was variable among species, ranging between 0.14 (Alpine salamanders) and 0.60 (cave salamanders). For all the species, abundance estimates obtained using N-mixture models showed limited differences with the ones obtained through capture-mark-recapture or removal sampling. The match was particularly accurate for cave salamanders in sites with limited abundance and for lizards, nevertheless non-incorporating heterogeneity of detection probability increased bias. N-mixture models provide reliable abundance estimates that are comparable with the ones of more traditional approaches, and offer additional advantages such as a smaller sampling effort and no need of manipulating individuals, which in turn reduces the risk of harming animals and spreading diseases.

The availability of data on the feeding habits of species of conservation value may be of great importance to develop analyses for both scientific and management purposes. Stomach flushing is a harmless technique that allowed us to collect extensive data on the feeding habits of six Hydromantes species. Here, we present two datasets originating from a three-year study performed in multiple seasons (spring and autumn) on 19 different populations of cave salamanders. The first dataset contains data of the stomach content of 1,250 salamanders, where 6,010 items were recognized; the second one reports the size of the intact prey items found in the stomachs. These datasets integrate considerably data already available on the diet of the European plethodontid salamanders, being also of potential use for large scale meta-analyses on amphibian diet.

Species can show strong variation of local abundance across their ranges. Recent analyses suggested that variation in abundance can be related to environmental suitability, as the highest abundances are often observed in populations living in the most suitable areas. However, there is limited information on the mechanisms through which variation in environmental suitability determines abundance. We analysed populations of the microendemic salamander Hydromantes flavus, and tested several hypotheses on potential relationships linking environmental suitability to population parameters. For multiple populations across the whole species range, we assessed suitability using species distribution models, and measured density, activity level, food intake and body condition index. In high-suitability sites, the density of salamanders was up to 30-times higher than in the least suitable ones. Variation in activity levels and population performance can explain such variation of abundance. In high-suitability sites, salamanders were active close to the surface, and showed a low frequency of empty stomachs. Furthermore, when taking into account seasonal variation, body condition was better in the most suitable sites. Our results show that the strong relationship between environmental suitability and population abundance can be mediated by the variation of parameters strongly linked to individual performance and fitness.

What are the long-term consequences of invasive species? After invasion, how long do ecosystems require to reach a new equilibrium? Answering these questions requires long-term, high-resolution data that are vanishingly rare. We combined the analysis of environmental DNA extracted from a lake sediment core, coprophilous fungi, and sedimentological analyses to reconstruct 600 years of ecosystem dynamics on a sub-Antarctic island and to identify the impact of invasive rabbits. Plant communities remained stable from AD 1400 until the 1940s, when the DNA of invasive rabbits was detected in sediments. Rabbit detection corresponded to abrupt changes of plant communities, with a continuous decline of a dominant plant species. Furthermore, erosion rate abruptly increased with rabbit abundance. Rabbit impacts were very fast and were stronger than the effects of climate change during the 20th century. Lake sediments can allow an integrated temporal analysis of ecosystems, revealing the impact of invasive species over time and improving our understanding of underlying mechanisms.

Leeches can parasitize many vertebrate taxa. In amphibians, leech parasitism often has potential detrimental effects including population decline. Most of studies on the host-parasite interactions involving leeches and amphibians focus on freshwater environments, while they are very scarce for terrestrial amphibians. In this work, we studied the relationship between the leech Batracobdella algira and the European terrestrial salamanders of the genus Hydromantes, identifying environmental features related to the presence of the leeches and their possible effects on the hosts. We performed observation throughout Sardinia (Italy), covering the distribution area of all Hydromantes species endemic to this island. From September 2015 to May 2017, we conducted >150 surveys in 26 underground environments, collecting data on 2629 salamanders and 131 leeches. Water hardness was the only environmental feature correlated with the presence of B. algira , linking this leech to active karstic systems. Leeches were more frequently parasitizing salamanders with large body size. Body Condition Index was not significantly different between parasitized and non-parasitized salamanders. Our study shows the importance of abiotic environmental features for host-parasite interactions, and poses new questions on complex interspecific interactions between this ectoparasite and amphibians.

Body mass is rarely recorded in amphibians, and other body measurements (e.g. snout to vent length, SVL) are generally collected instead. However, length measurements, when used as proxies of body mass in comparative analyses, are problematic if different taxa and morphotypes are included. We developed allometric relationships to derive body mass from SVL measurements. We fitted phylogenetic generalized least square models for frogs (Anura) and salamanders (Caudata) and for several families separately. We tested whether allometric relationships differed between species with different habitat preferences and between morphs in salamanders. Models were fitted with SVL-mass measurements for 88 frog and 42 salamander species. We assessed the predictive performance of the models by cross-validation. Overall, the models showed high explained variance and low forecasting errors. Models differed among semi-aquatic, terrestrial and arboreal frogs, and between paedomorphic and non-paedomorphic salamanders. Body mass estimates derived from our models allow for comparability of studies on multiple taxa and can be used for testing theories built upon evolutionary and ecological processes which are directly related to body mass., (© 2017 International Society of Zoological Sciences, Institute of Zoology/Chinese Academy of Sciences and John Wiley & Sons Australia, Ltd.)

The aim of this work was to evaluate the Ascidian Embryo Teratogenicity assay (AET) as new alternative invertebrate model to test the developmental effects of the co-exposure to ethanol and fluconazole. Ciona intestinalis embryos were exposed to the azolic fungicide fluconazole, (FLUCO, 7.8-250μM), to ethanol (Eth, 0.01-0.5%) and to their mixture (0.01% Eth+FLUCO 7.8-250μM) from neurula to larval stage. At the end of the exposure period, larvae were morphologically evaluated and benchmark analysis performed by using the PROAST modelling software. Both compounds were teratogenic in a concentration-related manner, particularly affecting the pigmented organs. The co-exposure to Eth enhanced the effects of FLUCO, the additive hypothesis was not rejected by the modelling. The results demonstrated that AET could be considered a good vertebrate-free alternative model for toxicological investigation in embryos., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Seasonality impacts species distributions through changes of the environmental conditions that affect the presence of individuals at a given place. Although the dynamics of cave microclimates are well known, only a few studies have evaluated the effects of such dynamics on non-strictly cave species. Here we assessed if species exploiting subterranean environments show changes in habitat occupation related to seasonal variation of cave microclimates. We surveyed 16 caves in central Italy every month for one year. Caves were subdivided into longitudinal sectors of three meters. In each sector we measured cave morphology and microclimatic features, assessed the occurrence of eight non-troglobitic taxa (orthopterans, spiders, gastropods and amphibians), and related species distribution to environmental features and sampling periods. The occurrence of most species was related to both cave morphology and microclimatic features. The survey month was the major factor determining the presence of species in cave sectors, indicating that cave-dwelling taxa show strong seasonality in activity and distribution. For multiple species, we detected interactions between sampling period and microclimatic features, suggesting that species may associate with different microhabitats throughout the year. The richest communities were found in sites with specific microclimates (i.e., high humidity, warm temperature and low light) but seasonality for species richness was strong as well, stressing the complexity of interactions between outdoor and subterranean environments., Competing Interests: The authors declare there are no competing interests.

Convergent adaptive evolution of species' ecological niches-i.e., the appearance of similar niches in independent lineages-is the result of natural selection acting on niche-related species traits ("traits" hereafter) and contrasts with neutral evolution [1-4]. Although trait convergences are recognized as being of importance at the species scale, we still know little about the impact of species convergence on the overall trait and niche structure of entire biotas at large spatial scales [5]. Here, we map the convergent evolution of four traits (diet, body mass, activity cycle, and foraging strata) for mammal species and assemblages (defined at 200 × 200 km resolution) at a global scale. Using data on the geographic distributions, traits, and phylogenetic relationships of species and by comparing observed patterns of trait β-diversity to evolutionary neutral expectations, we show that trait convergence is not restricted to particular lineages but scales up to entire assemblages (i.e., whole species communities). We find region-wide biota convergence in traits between regions with similar climates, particularly between Australia and other continents. Pairs of assemblages that show trait divergence often involves Arctic regions where rapid evolutionary changes occurred in response to extreme climatic constraints. By integrating both macroecological and macroevolutionary approaches into a single framework, our study quantifies the crucial role of evolutionary processes such as natural selection in the spatial distribution and structure of large-scale species assemblages., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Contents 924 I. 925 II. 925 III. 927 IV. 929 V. 930 VI. 930 VII. 931 VIII. 933 IX. 935 X. 936 XI. 938 938 References 938 SUMMARY: Recent advances in sequencing technologies now permit the analyses of plant DNA from fossil samples (ancient plant DNA, plant aDNA), and thus enable the molecular reconstruction of palaeofloras. Hitherto, ancient frozen soils have proved excellent in preserving DNA molecules, and have thus been the most commonly used source of plant aDNA. However, DNA from soil mainly represents taxa growing a few metres from the sampling point. Lakes have larger catchment areas and recent studies have suggested that plant aDNA from lake sediments is a more powerful tool for palaeofloristic reconstruction. Furthermore, lakes can be found globally in nearly all environments, and are therefore not limited to perennially frozen areas. Here, we review the latest approaches and methods for the study of plant aDNA from lake sediments and discuss the progress made up to the present. We argue that aDNA analyses add new and additional perspectives for the study of ancient plant populations and, in time, will provide higher taxonomic resolution and more precise estimation of abundance. Despite this, key questions and challenges remain for such plant aDNA studies. Finally, we provide guidelines on technical issues, including lake selection, and we suggest directions for future research on plant aDNA studies in lake sediments., (© 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.)

We examined how butterfly species richness is affected by human impact and elevation, and how species ranges are distributed along the elevational gradient (200-2700 m) in the Isère Department (French Alps). A total of 35,724 butterfly observations gathered in summer (May-September) between 1995 and 2015 were analyzed. The number of estimated species per 100-m elevational band was fitted to the elevational gradient using a generalized additive model. Estimations were also performed on a 500 m × 500 m grid at low altitude (200-500 m) to test for the human impact on species richness using generalized least squares regression models. Each species elevational range was plotted against the elevational gradient. Butterfly richness along the elevational gradient first increased (200-500 m) to reach a maximum of 150 species at 700 m and then remained nearly constant till a sharp decrease after 1900 m, suggesting that after some temperature threshold, only few specialized species can survive. At low elevation, urbanization and arable lands had a strongly negative impact on butterfly diversity, which was buffered by a positive effect of permanent crops. Butterfly diversity is exceptionally high (185 species) in this alpine department that represents less than 5% of the French territory and yet holds more than 70% of all the Rhopalocera species recorded in France. Both climate and habitat shape the distribution of species, with a negative effect of anthropization at low altitude and strong climatic constraints at high altitude.

The distribution of living organisms on Earth is spatially structured. Early biogeographers identified the existence of multiple zoogeographical regions, characterized by faunas with homogeneous composition that are separated by biogeographical boundaries. Yet, no study has deciphered the factors shaping the distributions of terrestrial biogeographical boundaries at the global scale. Here, using spatial regression analyses, we show that tectonic movements, sharp changes in climatic conditions and orographic barriers determine extant biogeographical boundaries. These factors lead to abrupt zoogeographical transitions when they act in concert, but their prominence varies across the globe. Clear differences exist among boundaries representing profound or shallow dissimilarities between faunas. Boundaries separating zoogeographical regions with limited divergence occur in areas with abrupt climatic transitions. In contrast, plate tectonics determine the separation between deeply divergent biogeographical realms, particularly in the Old World. Our study reveals the multiple drivers that have shaped the biogeographical regions of the world.