Telomere length and dynamics in Astyanax mexicanus cave and surface morphs.

Background: Telomeres are non-coding DNA repeats at the chromosome ends and their shortening is considered one of the major causes of aging. However, they also serve as a biomarker of environmental exposures and their length and attrition is affected by various stressors. In this study, we examined the average telomere length in Astyanax mexicanus, a species that has both surface-dwelling and cave-adapted populations. The cave morph descended from surface ancestors and adapted to a markedly different environment characterized by specific biotic and abiotic stressors, many of which are known to affect telomere length. Our objective was to explore whether telomere length differs between the two morphs and whether it serves as a biological marker of aging or correlates with the diverse environments the morphs are exposed to. Methods: We compared telomere length and shortening between laboratory-reared Pachón cavefish and Rio Choy surface fish of A. mexicanus across different tissues and ages. Results: Astyanax mexicanus surface fish exhibited longer average telomere length compared to cavefish. In addition, we did not observe telomere attrition in either cave or surface form as a result of aging in adults up to 9 years old, suggesting that efficient mechanisms prevent telomere-mediated senescence in laboratory stocks of this species, at least within this time frame. Our results suggest that telomere length in Astyanax may be considered a biomarker of environmental exposures. Cavefish may have evolved shorter and energetically less costly telomeres due to the absence of potential stressors known to affect surface species, such as predator pressure and ultra-violet radiation. This study provides the first insights into telomere dynamics in Astyanax morphs and suggests that shorter telomeres may have evolved as an adaptation to caves.

Groundwater is a hidden global keystone ecosystem.

Groundwater is a vital ecosystem of the global water cycle, hosting unique biodiversity and providing essential services to societies. Despite being the largest unfrozen freshwater resource, in a period of depletion by extraction and pollution, groundwater environments have been repeatedly overlooked in global biodiversity conservation agendas. Disregarding the importance of groundwater as an ecosystem ignores its critical role in preserving surface biomes. To foster timely global conservation of groundwater, we propose elevating the concept of keystone species into the realm of ecosystems, claiming groundwater as a keystone ecosystem that influences the integrity of many dependent ecosystems. Our global analysis shows that over half of land surface areas (52.6%) has a medium-to-high interaction with groundwater, reaching up to 74.9% when deserts and high mountains are excluded. We postulate that the intrinsic transboundary features of groundwater are critical for shifting perspectives towards more holistic approaches in aquatic ecology and beyond. Furthermore, we propose eight key themes to develop a science-policy integrated groundwater conservation agenda. Given ecosystems above and below the ground intersect at many levels, considering groundwater as an essential component of planetary health is pivotal to reduce biodiversity loss and buffer against climate change.

Recent Advances in the Behavioral Ecology of European Plethodontid Salamanders.

There is a recent growing interest in the study of evolutionary and behavioral ecology of amphibians. Among salamanders, Plethodontidae is the most speciose family, with more than 500 species, while in Europe, there are only 8 species, all belonging to the genus Speleomantes. European plethodontids recently received increasing attention with regard to the study of their natural history, ecology and behavior; however, the lack of standardized data, especially for the latter, hampers comparative analysis with the species from the New World. We here synthetized the recent advances in Speleomantes behavioral ecology, considering as a starting point the comprehensive monography of Lanza and colleagues published in 2006. We identified the behavioral categories that were investigated the most, but we also highlighted knowledge gaps and provided directions for future studies. By reviewing the scientific literature published within the period 2006-2022, we observed a significant increase in the number of published articles on Speleomantes behavior, overall obtaining 36 articles. Behavioral studies on Speleomantes focused mainly on trophic behavior (42%), and on intraspecific behavior (33%), while studies on pheromonal communication and interspecific behavioral interactions were lacking. In addition, most of the studies were observational (83%), while the experimental method was rarely used. After providing a synthesis of the current knowledge, we suggest some relevant topics that need to be considered in future research on the behavioral ecology of European plethodontids, highlighting the importance of a more integrative approach in which both field observations and planned experiments are used.

Electrifying Friedel-Crafts Intramolecular Alkylation toward 1,1-Disubstituted Tetrahydronaphthalenes.

In this work, we successfully employed electrochemical conditions to promote a Hofer-Moest, intramolecular Friedel-Crafts alkylation sequence. The reaction proceeds under mild conditions, employing carboxylic acids as starting materials. Notably, the electrochemical process performed in batch was adapted to a continuous flow electrolysis apparatus to provide a significant improvement. This catalyst-free, electrochemical approach produces an array of tetrahydronaphthalenes that could be used for API synthesis.

Inferring on Speleomantes Foraging Behavior from Gut Contents Examination.

We here provide the first comprehensive analysis and discussion on prey consumed by the European cave salamanders of the genus Speleomantes. Our study stems from the need to shed light on the still unknown foraging behavior adopted by Speleomantes cave salamanders. Starting from the published datasets on gut contents from all Speleomantes species (including hybrids), we here discuss additional information (i.e., species ecology, lower taxonomic level), which were systematically omitted from those data sets. We analyzed a data set consisting of 17,630 records from 49 categories of consumed prey recognized from gut contents of 2060 adults and juveniles Speleomantes. Flying prey accounted for more than 58% of the prey items, while elongated crawling prey accounted for no more than 16% of the diet within a single population. Among the total recognized prey items, only three can be surely ascribed to the group of strictly-cave species (i.e., troglobites), meaning that European cave salamanders mostly forage in surface environment, and therefore represent one of the major drivers of allochthonous organic matter in subterranean environments. Some of the consumed prey seemed to be aquatic, allowing us to hypothesize whether Speleomantes are able to catch prey from a shallow body water. Furthermore, European cave salamanders possess the ability to prey upon taxa characterized by particular anti-predator defenses, while morphological constraints seem to be the most important limit to prey consumption. For each specific case, we provide insights and propose hypotheses concerning the foraging behavior that need to be tested to properly understand the foraging behavior of this cryptic salamanders.

Monitoring of the Endangered Cave Salamander Speleomantes sarrabusensis.

In this study, we performed the first monitoring activities on one of the most endangered amphibians in Europe, the Sette Fratelli cave salamander Speleomantes sarrabusensis. The data presented here are derived from two monitoring activities aiming to assess the status and abundance of four populations of S. sarrabusensis. With the first monitoring, we surveyed the well-known population occurring within artificial springs during the period 2015-2018, providing monthly data on the number of active individuals. With the second monitoring performed during spring to early summer of 2022, we surveyed four populations at three time points (the one from artificial springs and three from forested areas) and we provided the first estimation of the populations' abundance. Furthermore, we analyzed for the first time the stomach contents from a population of S. sarrabusensis only occurring in forested environments. With our study, we provided the first information on the abundance of different populations of S. sarrabusensis, representing the starting point for future status assessments for this endangered species.

The trophic niche of subterranean populations of Speleomantes italicus.

The determination of a species trophic niche can clarify its functional role within a food web and how prey resources are used in relation with the spatial and temporal variability of environmental conditions. This information may result particularly useful for the implementation of conservation plans of endangered species having a cryptic behaviour or living in places difficult to be surveyed. Here we present the first long-term study on the trophic niche of the Italian cave salamander Speleomantes italicus, a strictly protected facultative cave species that seasonally exploits surface environments (e.g., forested areas) as well as both natural and artificial subterranean environments. We analysed the diet variation of six populations of S. italicus inhabiting natural caves, surveyed 24 times in a full year. During the surveys, all sampled individuals were subjected to stomach flushing and the ingested prey were identified and enumerated; furthermore, salamanders' body condition was also evaluated. The results of the analyses provided the first comprehensive, year-round assessment of the diet for a Speleomantes species. Remarkable divergences in terms of trophic niche and body condition were observed between the studied populations. We found a discrepancy in the foraging activity of the populations located in different areas; specifically, the individuals that experienced sub-optimal microclimatic conditions poorly performed in foraging. Furthermore, we found temporal and spatial variability in the body condition of individuals. Our study highlighted a remarkably high spatial and temporal divergence in the trophic habits of conspecific populations, a feature that may represent one of the major factors promoting the variability of multiple population traits.

The Trophic Niche of Two Sympatric Species of Salamanders (Plethodontidae and Salamandridae) from Italy.

The trophic niche of a species is one of the fundamental traits of species biology. The ideal trophic niche of a species is realized in the absence of interspecific competition, targeting the most profitable and easy-to-handle food resources. However, when a competitor is present, species adopt different strategies to reduce competition and promote coexistence. In this study, we assessed the potential mechanisms that allow the coexistence of two generalist salamanders: the Italian cave salamander (Speleomantes italicus) and the fire salamander (Salamandra salamandra). We surveyed, in April 2021, a forested area of Emilia-Romagna (Italy) during rainy nights. Analyzing the stomach contents of the captured individuals, we obtained information on the trophic niche of these two sympatric populations. Comparing our results with those of previous studies, we found that the two species did not modify their trophic niche, but that alternative mechanisms allowed their coexistence. Specifically, different prey preferences and predator metabolisms were likely the major factors allowing reduced competition between these two generalist predators.

A trait database and updated checklist for European subterranean spiders.

Species traits are an essential currency in ecology, evolution, biogeography, and conservation biology. However, trait databases are unavailable for most organisms, especially those living in difficult-to-access habitats such as caves and other subterranean ecosystems. We compiled an expert-curated trait database for subterranean spiders in Europe using both literature data (including grey literature published in many different languages) and direct morphological measurements whenever specimens were available to us. We started by updating the checklist of European subterranean spiders, now including 512 species across 20 families, of which at least 192 have been found uniquely in subterranean habitats. For each of these species, we compiled 64 traits. The trait database encompasses morphological measures, including several traits related to subterranean adaptation, and ecological traits referring to habitat preference, dispersal, and feeding strategies. By making these data freely available, we open up opportunities for exploring different research questions, from the quantification of functional dimensions of subterranean adaptation to the study of spatial patterns in functional diversity across European caves.

Global response of conservationists across mass media likely constrained bat persecution due to COVID-19.

Most people lack direct experience with wildlife and form their risk perception primarily on information provided by the media. The way the media frames news may substantially shape public risk perception, promoting or discouraging public tolerance towards wildlife. At the onset of the COVID-19 pandemic, bats were suggested as the most plausible reservoir of the virus, and this became a recurrent topic in media reports, potentially strengthening a negative view of this ecologically important group. We investigated how media framed bats and bat-associated diseases before and during the COVID-19 pandemic by assessing the content of 2651 online reports published across 26 countries, to understand how and how quickly worldwide media may have affected the perception of bats. We show that the overabundance of poorly contextualized reports on bat-associated diseases likely increased the persecution towards bats immediately after the COVID-19 outbreak. However, the subsequent interventions of different conservation communication initiatives allowed pro-conservation messages to resonate across the global media, likely stemming an increase in bat persecution. Our results highlight the modus operandi of the global media regarding topical biodiversity issues, which has broad implications for species conservation. Knowing how the media acts is pivotal for anticipating the propagation of (mis)information and negative feelings towards wildlife. Working together with journalists by engaging in dialogue and exchanging experiences should be central in future conservation management.

Environmental DNA of insects and springtails from caves reveals complex processes of eDNA transfer in soils.

Subterranean environments host a substantial amount of biodiversity, however assessing the distribution of species living underground is still extremely challenging. Environmental DNA (eDNA) metabarcoding is a powerful tool to estimate biodiversity in poorly known environments and has excellent performance for soil organisms. Here, we tested 1) whether eDNA metabarcoding from cave soils/sediments allows to successfully detect springtails (Hexapoda: Collembola) and insects (Hexapoda: Insecta); 2) whether eDNA mostly represents autochthonous (cave-dwelling) organisms or it also incorporates information from species living in surface environments; 3) whether eDNA detection probability changes across taxa with different ecology. Environmental DNA metabarcoding analyses detected a large number of Molecular Operational Taxonomic Units (MOTUs) for both insects and springtails. For springtails, detection probability was high, with a substantial proportion of hypogean species, suggesting that eDNA provides good information on the distribution of these organisms in caves. Conversely, for insects most of MOTUs represented taxa living outside caves, and the majority of them represented taxa/organisms living in freshwater environments (Ephemeroptera, Plecoptera and Trichoptera). The eDNA of freshwater insects was particularly abundant in deep sectors of caves, far from the entrance. Furthermore, average detection probability of insects was significantly lower than the one of springtails. This suggests that cave soils/sediments act as "conveyer belts of biodiversity information", possibly because percolating water lead to the accumulation of eDNA of organisms living in nearby areas. Cave soils hold a complex mix of autochthonous and allochthonous eDNA. eDNA provided unprecedented information on the understudied subterranean cave organisms; analyses of detection probability and occupancy can help teasing apart local eDNA from the eDNA representing spatially-integrated biodiversity for whole landscape.

Capture-mark-recapture data on the strictly protected Speleomantes italicus.

This data set collects capture-mark-recapture data, biometric data, and stomach contents of seven populations of the Italian cave salamander (Speleomantes italicus), one of the strictly protected European plethodontid species endemic to mainland Italy. We monitored six subterranean populations inside caves, and one fully epigean population living in a forest, surveying a total area of >5200 m2 . Data collection was performed 24 times throughout a year for each of the subterranean populations, and seven times in late winter-early spring for the epigean population. Salamanders were individually identified using two different marking methods: subcutaneous injection of visual implant elastomers (VIE) and photographic recognition of the dorsal pattern. Overall, the data set contains information on 1283 captured salamanders, corresponding to 783 different individuals and 500 recapture events. This type of data can be used to assess the species detection probability and to estimate the size of the populations, which are fundamental parameters for the assessment of its conservation status. Captured salamanders were weighed using a digital scale and photographed next to a reference ruler to perform post hoc measurements. This allows to assess the potential variation of the body condition of individuals through the time, and the potential divergences between conspecific populations. Furthermore, repeated measurements of recaptured individuals can allow to evaluate the seasonal growth rates of Speleomantes. Before their release, the salamanders underwent stomach flushing, a non-invasive technique that allows us to investigate the food residues in the salamanders' stomach. In 951 salamanders, we were able to recognize a total of 7077 consumed prey items belonging to 37 different prey categories (i.e., order level or lower), completing the information on the consumed prey for the entire Speleomantes genus. Data on consumed prey can be used to assess potential divergences between populations or between individuals of different ages/sexes, but also to assess the potential trophic specialization of individuals. The distinctiveness of this data set is that, by combining the capture-mark-recapture data with those on the diet of individuals, it allows to perform detailed studies on the consistency of individuals' food preference over time, an analysis that has never been performed on these salamanders. We release the data set into the public domain under Creative Commons Attribution 4.0 International license (CC-BY). When you use this data in your publication, we request that you cite this data paper; if you are using the whole data set related to the entire project, please cite all the related papers. If this data set will be an important part of the data analyzed in your study, you should consider discussing a collaboration with the data set contact person. If you plan to use data from the European plethodontid salamanders trophic niche project, please contact the contact person to find out if similar analyses are already underway or if unpublished updates are available.

Selective Binding and Redox-Activity on Parallel G-Quadruplexes by Pegylated Naphthalene Diimide-Copper Complexes.

G-quadruplexes (G4s) are higher-order supramolecular structures, biologically important in the regulation of many key processes. Among all, the recent discoveries relating to RNA-G4s, including their potential involvement as antiviral targets against COVID-19, have triggered the ever-increasing need to develop selective molecules able to interact with parallel G4s. Naphthalene diimides (NDIs) are widely exploited as G4 ligands, being able to induce and strongly stabilize these structures. Sometimes, a reversible NDI-G4 interaction is also associated with an irreversible one, due to the cleavage and/or modification of G4s by functional-NDIs. This is the case of NDI-Cu-DETA, a copper(II) complex able to cleave G4s in the closest proximity to the target binding site. Herein, we present two original Cu(II)-NDI complexes, inspired by NDI-Cu-DETA, differently functionalized with 2-(2-aminoethoxy)ethanol side-chains, to selectively drive redox-catalyzed activity towards parallel G4s. The selective interaction toward parallel G4 topology, controlled by the presence of 2-(2-aminoethoxy)ethanol side chains, was already firmly demonstrated by us using core-extended NDIs. In the present study, the presence of protonable moieties and the copper(II) cavity, increases the binding affinity and specificity of these two NDIs for a telomeric RNA-G4. Once defined the copper coordination relationship and binding constants by competition titrations, ability in G4 stabilization, and ROS-induced cleavage were analyzed. The propensity in the stabilization of parallel topology was highlighted for both of the new compounds HP2Cu and PE2Cu. The results obtained are particularly promising, paving the way for the development of new selective functional ligands for binding and destructuring parallel G4s.

Collecting eco-evolutionary data in the dark: Impediments to subterranean research and how to overcome them.

Caves and other subterranean habitats fulfill the requirements of experimental model systems to address general questions in ecology and evolution. Yet, the harsh working conditions of these environments and the uniqueness of the subterranean organisms have challenged most attempts to pursuit standardized research.Two main obstacles have synergistically hampered previous attempts. First, there is a habitat impediment related to the objective difficulties of exploring subterranean habitats and our inability to access the network of fissures that represents the elective habitat for the so-called "cave species." Second, there is a biological impediment illustrated by the rarity of most subterranean species and their low physiological tolerance, often limiting sample size and complicating laboratory experiments.We explore the advantages and disadvantages of four general experimental setups (in situ, quasi in situ, ex situ, and in silico) in the light of habitat and biological impediments. We also discuss the potential of indirect approaches to research. Furthermore, using bibliometric data, we provide a quantitative overview of the model organisms that scientists have exploited in the study of subterranean life.Our over-arching goal is to promote caves as model systems where one can perform standardized scientific research. This is important not only to achieve an in-depth understanding of the functioning of subterranean ecosystems but also to fully exploit their long-discussed potential in addressing general scientific questions with implications beyond the boundaries of this discipline.

Updating salamander datasets with phenotypic and stomach content information for two mainland Speleomantes.

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.

The post hoc measurement as a safe and reliable method to age and size plethodontid salamanders.

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 (SVLe) 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 SVLe in terms of data quality and in reducing the stress caused to wild animals.

Do Chinese cavefish show intraspecific variability in morphological traits?

Cavefishes represent one of the most bizarre and intriguing life forms inhabiting groundwater environments. One-third of the known cavefishes worldwide is endemic to China, and almost half of those belongs to a single genus, Sinocyclocheilus (Cypriniformes: Cyprinidae). Analyzing the morphometrics of three Sinocyclocheilus species, we aimed to assess whether variability among conspecific populations exists. We predict that populations inhabiting different subterranean habitats (shallow vs. deep) show divergences in specific morphological traits to better cope with the local ecological conditions. Our results showed that the populations showing bigger eyes and reduced humpback were those occurring close to the cave entrance (habitats with light and high food availability), while specimens with smaller eyes and increased humpback were collected from deeper groundwater areas (habitats laying in darkness with food scarcity). This explorative study paves the way for further researches aiming to collect novel data on Chinese cavefishes and highlights the usefulness of these species in evolutionary studies.

Photographic database of the European cave salamanders, genus Hydromantes.

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.

Interspecific and interpopulation variation in individual diet specialization: Do environmental factors have a role?

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.