Valeria Messina

In northern Italy, the ongoing spread of non‐native red‐swamp crayfish Procambarus clarkii keeps indirectly decimating native populations of white‐clawed crayfish Austropotamobius pallipes complex and is now also threatening local amphibians adapted to slow‐flowing, hilly streams at the borders of its distribution. We conducted a preliminary field investigation into crayfish differential effects on the growth of fire salamander ( Salamandra salamandra ) larvae. LMMs with orthogonal contrasts revealed that scaled mass index (SMI) was significantly higher for larvae living with native crayfish, while tail damage was mainly explained by SMI and missing limbs were too rare for statistical analysis. This may suggest stronger predation pressure or indirect effects on resource availability by P. clarkii , which could threaten previously unimpacted sites leading to unanticipated amphibians decline.

Mosquitoes are vectors of numerous pathogens that pose significant threats to human and animal health, making their monitoring crucial for effective disease control. Environmental DNA (eDNA)–based approaches enable the detection of multiple organisms in their habitats without the need for morphological identification. However, they require strict and extensive protocol optimization. In this study, we focused on the need for novel monitoring strategies, particularly ones capable of overcoming the limitations of traditional methods. Given the central role of water in the life cycle of mosquitoes, we explored whether eDNA collected from aquatic habitats could provide an effective tool for mosquito detection and taxonomic identification. Specifically, we aimed to evaluate whether an eDNA‐based approach can reliably detect multiple mosquito species and accurately discriminate among them under different community compositions. To this end, we developed and validated a protocol for an eDNA‐based detection and identification method for mosquito monitoring. First, we assessed its performance by conducting laboratory experiments on samples containing nine different combinations of larvae from five mosquito species. eDNA occupancy analysis revealed varying species‐specific detection probabilities, with Aedes albopictus showing the highest detection rate and Aedes koreicus the lowest. Overall detection probability was high at moderate to high larval densities (5 to 50 ind/L), whereas it decreased at 0.5 ind/L, indicating reduced sensitivity at very low larval concentrations. Then, we collected 165 water samples from three types of field locations and assessed the influence of ecological variables, including land use and site type, on mosquito detection probability. Our results indicate that eDNA represents a promising tool for mosquito monitoring, particularly in contexts where larval densities are not extremely low, and may support its future integration into vector surveillance and control program.

Ecologists and zoologists have worked for centuries to understand the spreading, extent, and regulation of biological rhythms among eukaryotes living in arhythmic ecosystems. Investigating whether the long‐standing hypothesis that biological clocks went lost in species inhabiting arhythmic environments can thus have broad implications in assessing evolutionary and ecological pathways allowing colonization of new and physiologically challenging environments. Subterranean habitats generally lack temporal variation in environmental conditions. However, chronobiological research in such communities remains hindered by methodological, ethical, and logistical constraints, particularly for field‐based studies. Here, we propose a flexible, noninvasive framework to assess temporal variation in the activity of animals using detection probability as a proxy for biological rhythms. Our approach is based on Bayesian occupancy and abundance models, which can incorporate quantitative (count), qualitative (presence/absence), or semiquantitative (abundance classes) data derived from repeated surveys. This framework is adaptable to a variety of taxa, monitoring strategies, and environmental conditions. It is particularly well‐suited for sensitive ecosystems, as it minimizes disturbance and allows for standardized, repeatable analysis of activity patterns. We illustrate the utility of the framework through three case studies ( Proteus anguinus Laurenti 1768, Niphargus sp. Schiødte 1849, and Spelaeomysis bottazzii Caroli 1924) encompassing different data types and showing its potential to detect rhythmicity even in stable subterranean environments. By facilitating robust in situ assessments of temporal activity, our method provides a novel and accessible tool for exploring biological rhythms in arhythmic environments, and can thus support the conservation of highly specialized, often vulnerable, fauna.

Springs represent ecotones between groundwater and surface freshwater habitats. Recent research suggested that springs can be more important than expected for stygobiont (i.e., adapted to live in groundwater) species, still information on habitat exploitation and activity of stygobionts in springs is far from complete. The aims of this study are: (i) to identify environmental factors promoting the exploitation of ecotone habitats by the stygobiont shrimp Troglocaris planinensis , commonly found in spring environments in northeastern Italy; and (ii) to experimentally evaluate whether this species exhibits differential behavioral responses to light and to subterranean and surface predator cues based on its habitat of origin (spring versus cave). From June 2020 to January 2025, we started multiple day and night surveys of T. planinensis in 64 springs of the Classic Karst (NE‐Italy). Each site has been characterized with respect to abiotic and biotic features. In the laboratory, shrimps from both cave and spring populations were tested to assess behavioral differences in response to light stimuli and predatory cues, as potential adaptations to the contrasting conditions of their respective habitats. In springs, T. planinensis density reached up to 116 shrimps/m ² , with significantly higher counts at night and lower densities at sites with greater fish predator abundance. Laboratory tests showed that predator cues, but not light exposure, influenced shrimp behavior regardless of their cave or spring origin. This study suggests that stygobiont crustaceans can represent a significant portion of biomass in surface waters and exploit these environments in response to changes in abiotic and biotic conditions and stimuli. However, further research is necessary to determine how stygobionts perceive surface conditions and how ecotonal pressures may drive adaptive shifts in typically groundwater‐dwelling animals.

Springs are ecotones between groundwater and surface water, important for a variety of both surficial and subterranean organisms. However, their use by amphibians has been poorly assessed. This is evident considering estavelles, typical ecotones functioning not only as spring but also as sinkholes. Here we aim to assess the use of estavelles by amphibians in the Classical Karst Region. From June 2020 to January 2025, we surveyed 61 springs, both during day and night. We visually recorded the occurrence of amphibians, along with abiotic and biotic features—including the presence of pikes (Esox cisalpinus), water flow level, drying events, and substratum heterogeneity. Forty-three springs exhibited estavelle-like hydrological behavior at least once. We identified five amphibian species. The use of springs was frequent for Proteus anguinus, Rana latastei, and Pelophylax sp. Amphibians’ occurrence was significantly linked to non-perennial estavelles characterized by low water flow, habitat heterogeneity, and absence of pike. Overall, reproductive activity in estavelles was limited; breeding likely occurs in nearby damp lentic habitats connected to the springs. Our results provide the first herpetological assessment of estavelle spring habitats in the Classical Karst Region, emphasizing their role as shelters for epigean species and feeding patches for stygobionts.

Detecting the trends of species and populations is fundamental to identifying taxa with high conservation priority. Unfortunately, long-term monitoring programs are challenging and often lacking. The Italian agile frog Rana latastei is endemic to Northern Italy and adjacent countries, is considered vulnerable by the IUCN, and is protected at the European level. However, quantitative estimates of its decline are extremely scarce. In this study, we document the trends in abundance and distribution of Rana latastei within Monza Park, which currently represents the area closer to the type locality of the species and holds unique genetic features. Wetlands within the park were monitored from 2000 to 2023; counts of egg clutches were taken as a measure of reproductive output and the abundance of breeding females. In 2000, the species occurred over a significant proportion of the park. Total abundance showed strong yearly variation but remained rather constant from 2000 to 2019. However, Rana latastei disappeared from the park around 2021 and was never detected in 2022–2023. The decline is probably related to the joint effect of multiple factors, including the conversion of breeding sites for farming, inappropriate water management, invasive alien species, and severe drought. The local extinction of Rana latastei occurred despite legal protection, highlighting the need for more effective and stringent tools for the conservation of European biodiversity.

Human dimensions in endangered species conservation are often overlooked. However, including human dimensions in conservation projects can help conservationists integrate social and ecological dimensions and investigate individual or society’s perspectives toward conservation efforts. The aim of this study was to investigate how visitors of a protected area perceived the conservation of the native white-clawed crayfish (Austropotamobius pallipes), a species recently brought to disappearance by the introduction of the spiny-cheek crayfish (Faxonius limosus). From August to October 2017, we distributed a questionnaire to park visitors, aiming to: (i) identify behaviors that could affect the conservation of the native crayfish after reintroduction; (ii) investigate if visitors’ awareness and knowledge of native freshwater crayfish may increase approval for conservation initiatives. Of the 290 respondents, most (73%) had never heard of alien species. The respondents’ inclination to be favorable to resource investment and alien crayfish eradication and to perform specific educational training was significantly related to their knowledge of crayfish features. Knowledge of freshwater crayfish features resulted in being a main determinant in affecting visitors’ attitudes towards conservation initiatives. These results confirm that visitors’ awareness can increase approval for conservation programs and the importance of investigating the proper human dimensions before establishing management actions for endangered species.

Detecting the trends of species and populations is fundamental to identify taxa with high conservation priority. Unfortunately, long-term monitoring programs are challenging and often lacking. The Italian agile frog Rana latastei is endemic to Northern Italy and adjacent countries, is considered vulnerable by the IUCN, and is protected at the European level. However, quantitative estimates of its decline are extremely scarce. In this study, we document the trends in abundance and distribution of Rana latastei within the Monza Park, which currently represents the area closer to the type locality of the species. Wetlands within the park were monitored from 2000 to 2023; counts of egg clutches were taken as a measure of reproductive output and of the abundance of breeding females. In 2000, the species occurred over a significant proportion of the park. Total abundance showed strong yearly variation but remained rather constant from 2000 to 2019. However, Rana latastei disappeared from the park around 2021 and was never detected in 2022-23. The decline is probably related to the joint effect of multiple factors, including the conversion of breeding sites for farming, inappropriate water management, invasive alien species, and severe drought. The local extinction of Rana latastei occurred despite legal protection, highlighting the need for more effective and stringent tools for the conservation of European biodiversity.