Claudio Ferrari

Aim: Freshwater rock pools are ephemeral and fragile habitats that support specialised animal taxa. While distributed worldwide, these habitats are usually neglected and overlooked. We used DNA metabarcoding and metaphylogeographic approaches to study inter and intraspecific tardigrade biodiversity to identify their biogeographic patterns to inform conservation measures. Location: Europe. Period: Present. Major Taxa Studied: Tardigrades. Methods: We conducted DNA metabarcoding targeting a fragment of the cytochrome oxidase subunit 1 gene of 163 freshwater rock pools from four sites in Sweden, Poland and Italy to analyse the effect of both short and long distance on their alpha diversity and genetic differentiation. Results: A total of 85 tardigrades OTUs (species‐level clusters) were detected, with the most prevalent taxa across all samples being species from the genus Ramazzottius . OTUs richness was mostly driven by a negative relationship with rock pool depth and site‐specific differences. For seven OTUs a phylogeographic analyses revealed a strong founder effect and a variable effect of geographic distance and climate on their population structures. Main Conclusions: The high biodiversity and presence of potential specialised taxa highlight the conservation value of freshwater rock pools. While some tardigrade species may be able to easily disperse and have a large geographic range, other species show isolation by distance patterns and their genetic diversity may be negatively affected by habitat loss. Our results call for a bigger effort in identifying, mapping, characterising and protecting freshwater rock pools as underlooked but important habitats.

The Italian wolf Canis lupus italicus faced near extinction in the 19th and early 20th centuries due to extensive persecution, habitat degradation, and prey depletion. Over the past 50 years, conservation efforts, legal protection, and habitat recovery have supported a resurgence, with recent estimates placing the population at approximately 3,300 individuals. Wolves have expanded into lowlands, plains, and coastal areas, including the highly anthropised Po Plain. This expansion intensifies human-wildlife conflicts, particularly in livestock farming and hunting. The invasive coypu (Myocastor coypus) and eastern cottontail (Sylvilagus floridanus) have inflicted considerable ecological and economic damage in northern Italy. These populations are poorly controlled due to their ecological plasticity and lack of natural predators. Given the adaptability of wolves and their opportunistic diet, this study investigates whether they could become a significant prey species for wolves in plains environments, potentially contributing to their control while mitigating livestock predation. Through genetic analysis, scat morphology, and camera trapping, this study confirms the presence of coypu in the diet of a wolf pack in the Torrile and Trecasali Natural Reserve. These findings highlight the potential ecological role of wolves as indirect allies in managing invasive species, which could help alleviate economic and physical damage to agriculture and riverbanks, fostering more positive public attitudes and greater acceptance of the species.

The European endemic barbels represent important bioindicators of river quality and are nowadays threatened by changing environmental conditions and hybridization with the invasive alien Barbus barbus. It is therefore fundamental to investigate interactions among species and adaptability to climate changes in protected areas of Northern Apennines. An investigation was carried out considering 248 barbel samples that were analysed for Cytb mitochondrial DNA and 192 at 10 microsatellite loci, to delineate the distribution and population structure of the two native species as well as the impact of invasive B. barbus inside 15 sites of the Natura 2000 network. The complex distribution of the native barbel species was highlighted, together with a significant genetic structure emerging in different populations. Only one site revealed a “pure” population of B. caninus while the other ones showed a high level of hybridization between the different barbel species. For the B. plebejus two “genetically pure” populations were found in the hill‐mountain sector, while the hybridization level resulted in increasing in the lowest altitudinal stretch of watercourses with a consistent contribution driven by B. barbus. We herein present the first evidence of B. barbus introgression along an altitudinal gradient, carried from the lowland water course to mountain stretches driven by B. plebejus migration. B. plebejus is the species that can act as vicariant organism able to transfer the B. barbus genome from the plain habitats into B. caninus genome of the higher altitude waterstreams, as a consequence of habitat shifts due to climate changes and anthropogenic acitivities.

Background: Macrophytes may modify benthic biodiversity and biogeochemistry via radial oxygen loss from roots. This condition contrasts sediments anoxia, allows roots respiration, and facilitates aerobic microbial communities and processes in the rhizosphere. Simultaneously, the rhizosphere can stimulate anaerobic microorganisms and processes via exudates or by favoring the build-up of electron acceptors as nitrate. As eutrophication often results in organic enrichment in sediments and large internal nutrients recycling, an interesting research question is to investigate whether plants maintain the capacity to stimulate aerobic or anaerobic microbial communities and processes also under elevated organic pollution. Methods: A manipulative experiment was carried out under laboratory-controlled conditions. Microcosms containing bare sediments and sediments transplanted with the macrophyte Vallisneria spiralis L. were created. The effect of the plant was investigated on sediments with moderate (8%) and elevated (21%) organic matter content, after an acclimatization period of 30 days. Chemical and physical parameters, microbial community composition and the potential rates of nitrification, denitrification and nitrate ammonification were measured at two different depths (0–1 and 1–5 cm) after the acclimatization period to evaluate the role of roots. Results: Vallisneria spiralis grew and assimilated pore water nutrients at the two organic matter levels and vegetated sediments had always nutrient-depleted porewaters as compared to bare sediments. Nitrifying microbes had a lower relative abundance and diversity compared to denitrifying bacteria. However, regardless of the organic content, in vegetated sediments nitrifiers were detected in deeper horizons as compared to bare sediments, where nitrification was confined near the surface. In contrast, potential denitrification rates were not affected by the presence of roots, but probably regulated by the presence of nitrate and by root-dependent nitrification. Potential nitrate ammonification rates were always much lower (< 3%) than potential denitrification rates. Conclusions: Vallisneria spiralis affects N-related microbial diversity and biogeochemistry at moderate and elevated organic matter content, smoothing bottom water–pore water chemical gradients and stimulating nitrification and nitrogen loss via denitrification. These results suggest the possibility to deploy V. spiralis as a nature-based solution to counteract eutrophication in freshwater systems impacted by high loads of organic matter, for example, downstream of wastewater treatment plants.

The recent increase in international fish trade leads to the need for improving the traceability of fishery products. In relation to this, consistent monitoring of the production chain focusing on technological developments, handling, processing and distribution via global networks is necessary. Molecular barcoding has therefore been suggested as the gold standard in seafood species traceability and labelling. This review describes the DNA barcoding methodology for preventing food fraud and adulteration in fish. In particular, attention has been focused on the application of molecular techniques to determine the identity and authenticity of fish products, to discriminate the presence of different species in processed seafood and to characterize raw materials undergoing food industry processes. In this regard, we herein present a large number of studies performed in different countries, showing the most reliable DNA barcodes for species identification based on both mitochondrial (COI, cytb, 16S rDNA and 12S rDNA) and nuclear genes. Results are discussed considering the advantages and disadvantages of the different techniques in relation to different scientific issues. Special regard has been dedicated to a dual approach referring to both the consumer’s health and the conservation of threatened species, with a special focus on the feasibility of the different genetic and genomic approaches in relation to both scientific objectives and permissible costs to obtain reliable traceability.

The biodiversity protection and monitorning is one of main goals of natural history musems worldwide. Conservation issues are particularly important for freshwater fish which are one of the most threatened taxa for the consequences of climate change and human activies. In Mozambique freshwater rivers are poorly explored and the impact of aquaculture and human activities on local biodiversity in almost unknown. Here we propose the barcoding analysis of cytochrome c oxidase I (COI) mitochrondrial DNA of 41 frehswater fishes catched in four rivers of southern Mozambique and 53 from a museum collection. As evidence of previous knowledge gaps, barcoding results revealed twenty new haplotypes described for the first time in the taxa Cichlidae and Amphilidae. From a methodological point of view, the barcoding approach demonstrated a critical point connected to the requested 650 bp length of amplified sequences. In fact, high weight genomic DNA is unattainable from museum samples and also in wildlife samples collected in pristine rivers. For this reason we furtherly tested the efficiency of DNA mini-barcoding analysis for 53 fish from a museum collection. The Mini-barcode method retrieved 56.6% of sequences successfully analyzed versus 3% of barcoding. The high performance of this thecniques is discussed in relation to biodiversity monitoring and to fill the taxonomy gaps in museum collections.

Rock pools are ephemeral freshwater habitats characterized by their small size, well-defined boundaries, and periodic desiccation, making them ideal model systems to answer numerous ecological questions. Although there are numerous studies on rock pool fauna around the world, tardigrades have only rarely been recorded. We conducted the first tardigrade-focused study on rock pools by quantitatively extracting and classifying them from rock pools in the Italian Apennines. Rock pools were divided into three types, based on maximum duration of their inundation period. Following the patterns usually observed with rock pool invertebrates, we tested the hypothesis that desiccation has a negative effect on prevalence, abundance, and diversity of tardigrades, and thus could alter the trophic structure of their communities. In contrast to what is commonly found for other animal groups in the same type of habitat, we found that tardigrades were more diverse and prevalent in shallower, more desiccation prone, rock pools. Moreover, the trophic structure of tardigrade communities was different among the different pool types. Lastly, we also provide DNA barcodes of the most commonly found taxa. Ultimately, our study demonstrates that tardigrade communities in rock pools provide a valuable model system for the study of abiotic factors influencing meiofauna communities.

Climate change and water abstraction may change stream flow from perennial into intermittent lotic systems, modifying their abiotic and biotic benthic environment and impacting ecosystem processes such as nutrient turnover. We conducted a microcosm experiment to investigate the interactive effect of water intermittency, macrofauna and leaf size (Populus nigra leaves) on nutrient mineralization and recycling. Leaf disks (1 or 5 cm diameter) were incubated for 40 days with or without the leaf-consumer, Potamophylax cingulatus larvae (Trichoptera, Limnephilidae) and with or without an intervening, 10-days simulation of stream drying and subsequent rewetting. Nutrient fluxes, residual leaf biomass and leaf elemental composition were measured to evaluate how intermittency, macrofauna and leaf size affect organic matter mineralization rates and stoichiometry. Results suggest that drying slows decomposition rates, impacting both the microbial and setting to zero macrofauna activities. The presence of macrofauna increases mineralization and nutrient (C, N and P) regeneration rates. Our findings also suggest that leaf disks with higher diameter display higher microbial activity and NH₄⁺ regeneration. During the experiment, the C:N:P ratios of residual litter changed, as the leaf material became enriched with N and P. Our study suggests that increasingly frequent dry events might slow mineralization rates and downstream nutrient transport.

Presence of Danube barbel Barbus balcanicus was investigated at the westernmost portion of its distribution area (Italian portion of the Isonzo River Basin). Genetic analyses carried out on specimens collected in different watercourses confirmed B. balcanicus presence in the same locations where Barbus caninus was erroneously classified in the past. More precisely, 4 Cyt b haplotypes belonging to the Balkanic species were described, three of which were evidenced for the first time. Analysis of meristic characters also confirmed genetic results and completed the taxonomical assessment. At the same time, a target population was studied to deepen knowledge about the species ecology. In this case, 120 specimens belonging to five age classes (from 0+ to 4+) were collected with monthly frequency in Barbucina Creek, representing the model watercourse inhabited by the species in the study area. Regression between total weight (W) and standard length (S_L) did not differ significantly between males and females, therefore not displaying sexual dimorphism. S_L values showed wide ranges, often overlapping among age classes. Mean condition factor (Kmean) decreased significantly with growth and age, as small individuals were in better nutritional condition. From an ecological point of view, biometric parameters seemed to be affected by habitat conditions and, specifically, to the limited space of the creeks, rather than by trophic conditions. Finally, trends of gonadosomatic index (GSI) showed that B. balcanicus reproductive period stretches between April and late May/early June. Results indicated that sexual maturity is reached at 1+ age, corresponding to a mean standard length equal to 5.50 ± 0.66 for female and 5.71 ± 0.53 for male breeders. In conclusion, experimental results highlighted the presence and adaptability of this “forgotten” species of the Italian fish fauna, which certainly will deserve attention in a future amendment of the Italian IUCN Red List of vertebrates.