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Local adaptation and phenotypic plasticity can lead to environment-related morphological and genetic variations in freshwater fish. Studying the responses of fish to environmental changes is crucial to understand their vulnerability to human-induced changes. Here, we used a latitudinal gradient as a proxy for past and present environmental factors and tested its influences on both genetic and morphological patterns. We selected as a suitable biogeographic model, the barbels, which inhabit 17 Adriatic basins of the central-southern Italian Peninsula, and explored association among attributes from genetic, morphological, and environmental analyses. The analysis of the mitochondrial DNA control region evidenced a southward significant increase in the number of private haplotypes, supporting the isolation of the southernmost populations related to the Mio-Pleistocene events. In contrast, morphology was mainly affected by changes in the present environmental conditions. Particularly, the number of scales and fish coloration were clearly associated to latitude, and thus thermal and hydrological conditions. Other morphometric and functional traits varied under the selective pressure of other environmental factors like elevation and distance from headwater. These results highlight the sensitivity of barbels to climate changes, which can serve as a basis for future eco-evolutionary and conservation studies.

Freshwater bivalves are highly threatened and globally declining due to multiple anthropogenic impacts, making them important conservation targets. Because conservation policies and actions generally occur at the species level, accurate species identification and delimitation is critical. A recent phylogenetic study of Italian mussel populations revalidated an Anodonta species bringing the number of known European Anodontini from three to four species. The current study contributes to the clarification of the taxonomy and systematics of European Anodontini, using a combination of molecular, morphological and anatomical data, and constructs phylogenies based on complete mitogenomes. A redescription of A. exulcerata and a comparative analysis of morphological and anatomical characters with respect to the other two species of Anodonta present in the area are provided. No reliable diagnostic character has emerged from comparative analysis of the morphometric characters of 109 specimens from 16 sites across the Italian peninsula. In fact, the discriminant analysis resulted in a greater probability of correct assignment to the site of origin than to the species. This confirms the difficulties of an uncritical application of visual characters for the delimitation of species, especially for Anodontinae. This research was developed under ConBiomics: the missing approach for the conservation of freshwater bivalves project Nº NORTE-01-0145-FEDER-030286, cofinanced by COMPETE 2020, Portugal 2020 and the European Union through the ERDF - European Regional Development Fund and by FCT - Fundação para a Ciência e a Tecnologia, through national funds (UID/Multi/04423/2019). FCT also supported Manuel Lopes-Lima (SFRH/BD/115728/2016). info:eu-repo/semantics/publishedVersion

The Italian peninsula is a biodiversity hotspot, with its freshwater fish fauna characterized by high levels of local endemism. Two endemic fluvio‐lacustrine fishes of the genus Barbus (barbel, family Cyprinidae) have allopatric distributions in the Tyrrhenian and Adriatic basins of Italy. Barbus plebejus inhabits the mid‐ to northern Adriatic basins, while B. tyberinus is widespread in all central‐northern basins draining into the Tyrrhenian Sea. For basins in Southern Italy draining into the southern parts of these seas, there remains a knowledge gap on their barbel populations due to no previous genetic and morphological studies, despite their apparent biogeographic isolation. Correspondingly, this study quantified the presence and distribution of barbels in the Adriatic and Tyrrhenian basins of Southern Italy through genetic and morphological analyses of 197 fish sampled across eight populations. Testing of how local isolation has influenced the evolution and persistence of these populations was completed by examining sequence variation at two mitochondrial loci (cytochrome b and D‐loop) and performing geometric morphometric analyses of body shape, plus measuring 11 morphometric and meristic characters. Phylogenetic and morphological analyses revealed the presence of two genetically distinct lineages that differed significantly from adjacent B. tyberinus and B. plebejus populations. These two new taxa, here described as SI1 and SI2 Barbus lineages, are highly structured and reflect a complex mosaic biogeographic pattern that is strongly associated with the underlying hydrographical scenarios of the basins. The geographic isolation of these basins thus has high evolutionary importance that has to be considered for maintaining endemism., In Southern Italy, apparent biogeographic isolation may have influenced barbel evolutionary structure. Through genetic and morphological analyses, we quantified the presence and distribution of barbels in Southern Italy describing two new taxa, highly structured and reflecting a complex mosaic biogeographic pattern that is strongly associated with the underlying hydrographical scenarios of the basins.

Invasive alien species are a major driver of biodiversity loss, with their impacts potentially more intense when complexes of cryptic species are involved. In freshwaters, the anthropogenic manipulation of fish communities has resulted in altered fish communities, and in Europe has increased the complexity of Phoxinus species assemblages. Here, we investigated the Phoxinus communities of the westernmost part of the Po river basin, where adjacent freshwater ecosystems (Alpine high-altitude lakes and lowland streams) are representative of different management strategies (i.e. manipulated fish communities via stocking in Alpine lakes vs. natural populations in streams). We tested the genetic composition of the cryptic Phoxinus populations inhabiting these waters, as the species are morphologically indistinct. Sequences of the mitochondrial cytochrome oxidase I (COI) were obtained from 239 specimens, with the results indicating that 17 Alpine high-altitude lakes are now populated by a complex of Phoxinus species, comprising P. septimaniae (native to the Mediterranean area of France), P. csikii (native to the Central Balkans) and P. lumaireul (native to the North Adriatic Sea basins). Their introduction resulted from their use as angling live baits. Minnow populations in lowland streams were primarily comprised of native P. lumaireul, with only a single P. csikii specimen detected. While nuclear sequences of the recombination activating gene 1 (RAG1) marker were not useful for tracking the presence of alien alleles in these stream populations, the COI data emphasised the importance of using molecular tools to investigate cryptic species complexes that have been modified by anthropogenic activities.

The reintroduction of the extinct beluga sturgeon (Huso huso L.), an anadromous species with economic and traditional relevance, is a priority in next conservation strategies in Northern Italy. The EU-LIFE NATURA project aims to reintroduce the beluga sturgeon in the Po River basin through a captive breeding program. Critical requirements for the success of the program are river connectivity and knowledge of genetic diversity of the selected broodstocks to ensure self-sustainability of reintroduced populations. Here, the four broodstocks used for the reintroduction of beluga sturgeon have been genetically screened, genotyping 13 loci and sequencing mitochondrial DNA cytochrome b (Cyt b) gene and the entire mitochondrial DNA control region (D-Loop). The four broodstocks showed a medium-high level of nuclear genetic variability and the presence of two sub-populations, evidencing a total level of inbreeding coefficients able to sustain the good potential as future breeders. Mitochondrial analyses showed a genetic variability comparable to wild populations, further strengthening the positive potential of the investigated broodstock. Therefore, this study, showed how the degree of genetic diversity found within the four broodstocks used for H. huso reintroduction in the Po River basin could be suitable to ensure the success of the program, avoiding the inbreeding depression associated with founder effect and captive breeding.

Invasions of alien fishes can result in considerable consequences for native biodiversity, including local extinctions of native species through genetic introgression. In Italy, the alien European barbel Barbus barbus was first detected in 1994. It has since undergone range expansion, raising conservation concerns on their impacts on endemic Barbus species, including Barbus plebejus and Barbus tyberinus. Here, the genetic and phenotypic consequences of B. barbus invasion in the Tyrrhenian and Adriatic basins of central Italy were assessed by comparing ‘invaded’ with ‘uninvaded’ river sections that remain free of B. barbus due to barriers preventing their upstream dispersal. In both basins, uninvaded sites were confirmed as B. barbus free, but the endemic populations had low genetic variability. In the invaded sections, haplotype and nucleotide diversity was relatively high, with introgression skewed towards B. barbus genes, with the barbel populations comprising of only 4% and 23% of pure B. tyberinus and B. plebejus respectively. Relatively high morphological differentiation was apparent between pure B. tyberinus and hybrid forms, whilst differences were less apparent between pure B. plebejus and their hybrid forms. Thus, the endemic Barbus species only persist in areas that remain free of invasive B. barbus, with this only due to river structures that impede their upstream movements. As these structures also limit the effective population size of the endemic species, conservation plans must reconcile B. barbus dispersal prevention with the need to increase the population connectivity of the endemics.

Two fluviolacustrine lineages (SI1 Barbus and SI2 Barbus) of the Barbus genus have been recently detected in the Apulia-Campania ichthyogeographic district (southern Italy). The aim of this study was to determine the taxonomic status of these lineages by comparing them with the two already-established Italian fluviolacustrine species Barbus plebejus and Barbus tyberinus through a more in-depth molecular and morphological investigation. Genetic analyses were performed on both mitochondrial (cytochrome b) and nuclear (growth hormone paralog 2) DNA markers, and morphological analyses were carried out on specific age classes and purebred populations. Molecular analyses detected four evolutionary lineages at the mitochondrial level, whereas the nuclear data set highlighted the strict evolutionary relation between B. plebejus sensu stricto and the new lineages, converged in the B. plebejus complex clade. The morphological analyses allowed us to discriminate SI1 Barbus and SI2 Barbus from both B. plebejus and B. tyberinus. The new taxa could be discriminated by the greatest maximum body height and the longest pre-orbital distance, respectively. Both the new lineages have longer ventral and pectoral fins than B. plebejus and B. tyberinus, a larger caudal fin than B. tyberinus and a lower number of scales along the lateral line than B. plebejus. Both molecular and morphological results suggested the two southern Italian lineages could be considered as distinct endemic species: the formal description of the new species Barbus samniticus sp. nov. (i.e., SI1 Barbus) and the revalidation of Barbus fucini Costa, 1853 (i.e., SI2 Barbus) were thus proposed, and, for both species, molecular and morphological diagnosis were provided.

© 2019 Royal Swedish Academy of Sciences Italian freshwaters are highly biodiverse, with species present including the native fishes Barbus plebejus and Barbus tyberinus that are threatened by habitat alteration, fish stocking and invasive fishes, especially European barbel Barbus barbus. In central Italy, native fluvio-lacustrine barbels are mainly allopatric and so provide an excellent natural system to evaluate the permeability of the Apennine Mountains. Here, the morphologic and genetic distinctiveness was determined for 611 Barbus fishes collected along the Padany–Venetian (Adriatic basins; PV) and Tuscany–Latium (Tyrrhenian basins; TL) districts. Analyses of morphological traits and mitochondrial DNA sequence data explored the natural and anthropogenic factors that have shaped their distribution ranges. Over 100 alien B. barbus were recorded in the Tiber basin (TL district) and Metauro basin (PV district). Comparisons of genetic and morphometric data revealed that morphometric data could identify alien B. barbus from native Barbus, but could not differentiate between B. tyberinus and B. plebejus. Genetic analyses revealed ~50 D-loop mtDNA haplotypes and identified a distinct Barbus lineage present only in the Vomano River at the southern boundary of PV district. Demographic expansion and molecular variance analyses revealed a lack of geographic structuring across the sampling regions. While the contemporary B. plebejus distribution has been driven primarily by anthropogenic fish translocations, the dispersal of B. tyberinus has been via natural dispersion, including their crossing of the Apennine Mountains via temporary river connectivity. The results also revealed that the Barbus fishes of the mid-Adriatic region of Europe have a complex pattern of local endemism. To conserve these patterns of genetic uniqueness, especially in the mid-Adriatic basins, Barbus fishes should be managed by treating them as unique evolutionary units and ceasing translocations of all Barbus fishes between river basins.

The potential effects of climate change on the distribution of Mediterranean plants at their peripheral range make them extremely vulnerable to environmental fluctuations. The northern-most Italian population of Cistus albidus provides an outstanding case for study, as it grows at the extreme northern edge of its range and because phenological data have been available since the early XIX century. In the period 2012–2017, we analysed the population demography and spatial structure, growth and flowering phenology, and genetic variability to understand the history, current state and possible evolution of this population. Lastly, the population data were tested for association with environmental factors. The population studied consists of plants on average 10 years old, showing the highest growth rates in spring and autumn. Anthesis starts in April and ends in early June and is significantly dependent on temperature. High levels of genetic variability are present (average He = 0.471), with heterozygote excess detected at four loci out of five. Bayesian analysis does not support any genetic structuring of the population. The aggregated pattern in the distribution of individuals and their phenological traits, together with the absence of genetic structuring, seem to suggest that this extreme geographical population is not currently endangered.

Cryptobiosis is a reversible ametabolic state of life characterized by the ceasing of all metabolic processes, allowing survival of periods of intense adverse conditions. Here we show that 1) entire moss individuals, dated by 14C, survived through cryptobiosis during six centuries of cold-based glacier burial in Antarctica, 2) after re-exposure due to glacier retreat, instead of dying (due to high rates of respiration supporting repair processes), at least some of these mosses were able to return to a metabolically active state and remain alive. Moss survival was assessed through growth experiments and, for the first time, through vitality measurements. Future investigations on the genetic pathways involved in cryptobiosis and the subsequent recovery mechanisms will provide key information on their applicability to other systematic groups, with implications for fields as divergent as medicine, biodiversity conservation, agriculture and space exploration.

Freshwater mussels of the order Unionida have been dramatically declining globally. Despite their ecological importance, conservation of these animals has been hindered by unresolved taxonomy and a lack of data on the distribution and status of populations, especially in southern Europe. Although the Italian Peninsula has been noted as a centre of endemism and one of the major refugia of the glacial ages for several taxa, few studies have been performed on the genetic diversity of Unionida. Most importantly, the taxonomic status of several freshwater mussel populations of the Italian Peninsula is still unresolved. Here we present the first comprehensive dataset for the Unionida of the region spanning Italy and the coastal Croatian region (west of the Dinaric Alps). In total, 191 specimens were collected (85 Anodonta, 64 Unio, 17 Microcondylaea bonellii and 25 Sinanodonta woodiana) from 34 sites across the Italian Peninsula and coastal Croatian river basins for molecular identification (COI, 16S and 28S). Genetic analyses were performed to understand major phylogenetic and phylogeographic patterns. Seven species were detected: three Anodonta species (A. anatina, A. cygnea and A. exulcerata), two Unio species (U. mancus and U. elongatulus), Microcondylaea bonellii, and the invasive Sinanodonta woodiana. The presence of three endemic species (A. exulcerata, U. elongatulus and M. bonellii) confirms the importance of the region as a centre of endemism for freshwater mussels. The Apennine Mountains act as an important biogeographic barrier., This work was partially supported by the Portuguese Foundation for Science and Technology (FCT) under grants SFRH/BPD/108445/2015 (EF) and SFRH/BD/115728/2016 (MLL). Cindy Bogan reviewed a draft of this manuscript. We are grateful to the two anonymous reviewers for their helpful remarks and suggestions that improved the quality of the manuscript; and to the Go¨teborg Natural History Museum for the loan of Swedish tissue samples., info:eu-repo/semantics/publishedVersion

The Lesser Flamingo Phoeniconaias minor is a nomadic species, which inhabits shallow alkaline lakes and pans in sub-Saharan Africa and southern Asia. The extent of genetic diversity and the degree of differentiation within and among populations are important factors to determine in order to help manage and conserve the species, categorised as Near Threatened by the IUCN. This study provides an assessment of the population structure of the two largest African populations of P. minor by genotyping six microsatellite loci from 30 individuals sampled on Lake Bogoria (Kenya) and 11 individuals from Makgadikgadi Sua Pan (Botswana). The alleles detected per locus ranged from four to 13. The Lake Bogoria population harboured 15 specific alleles, whereas the Makgadikgadi Sua Pan population only six alleles. Moderate genetic diversity (He = 0.64–0.69) was comparable with populations that have not suffered from demographic bottlenecks or inbreeding. The populations are genetically similar with little differentiation...

� Background and Aims Ulmus minor has been severely affected by Dutch elm disease (DED). The introduction into Europe of the exotic Ulmus pumila, highly tolerant to DED, has resulted in it widely replacing native U. minor populations. Morphological and genetic evidence of hybridization has been reported, and thus there is a need for assessment of interspecific gene flow patterns in natural populations. This work therefore aimed at studying pollen gene flow in a remnant U. minor stand surrounded by trees of both species scattered across an agricultural landscape. � Methods All trees from a small natural stand (350 in number) and the surrounding agricultural area within a 5-km radius (89) were genotyped at six microsatellite loci. Trees were morphologically characterized as U. minor, U. pumila or intermediate phenotypes, and morphological identification was compared with Bayesian clustering of genotypes. For paternity analysis, seeds were collected in two consecutive years from 20 and 28 mother trees. Maximum likelihood paternity assignment was used to elucidate intra- and interspecific gene flow patterns. � Key Results Genetic structure analyses indicated the presence of two genetic clusters only partially matching the morphological identification. The paternity analysis results were consistent between the two consecutive years of sampling and showed high pollen immigration rates (� 0� 80) and mean pollination distances (� 3 km), and a skewed distribution of reproductive success. Few intercluster pollinations and putative hybrid individuals were found. � Conclusions Pollen gene flow is not impeded in the fragmented agricultural landscape investigated. High pollen immigration and extensive pollen dispersal distances are probably counteracting the potential loss of genetic variation caused by isolation. Some evidence was also found that U. minor and U. pumila can hybridize when in sympatry. Although hybridization might have beneficial effects on both species, remnant U. minor populations represent a valuable source of genetic diversity that needs to be preserved.

The expression profile of genes involved in oxidative stress, metal toxicity, apoptosis/cell cycle, and protein folding was investigated in liver and testis of Sprague-Dawley rats at different time intervals after i.t. instillation of AgNP (20 nm, 50 μg/rat). At 7 days, selective changes in the expression of genes encoding oxidative stress-related enzymes, namely Gpx1, SOD, FMO2 and GAPDH were observed in hepatic and testicular tissues. Other genes implicated in oxidative stress (Txnrd1, Gss, Gsr), metal toxicity (Mt1), apoptosis/cell cycle (casp3, p53), and protein-folding processes (Hsp70) were not modified. Gene expression was modulated by AgNPs in a tissue- and time-dependent manner. In particular, SOD was up-regulated in both tissues, but significant Gpx1, FMO2, and GAPDH overexpression was seen in testes only. No gene expression changes were seen in both tissues 28 days post-instillation. Unlike AgNPs, pulmonary treatment with AgNO3 (7 μg/rat), did not cause gene expression changes in both tissues at both time points studied, suggesting that dissimilar mechanisms are implicated in toxicity and/or biokinetics of nanoparticulate and ionic silver. The results demonstrate subtle systemic changes involving selected oxidative stress-related genes in the liver and testis of animals exposed by pulmonary route to a low dose of AgNPs. These effects were apparently reversible as changes were observed at day 7 but not day 28. Recovery could possibly reflect either compensatory mechanism contrasting the initial toxicogenomic response to AgNPs or the silver removal from the tested organs. These findings may be of toxicological relevance in relation to possible health risks associated with occupational or consumer exposure to nanosilver.

Alcolapia is a minor genus of small-bodied, polymorphic cichlids inhabiting the lagoons and hot springs surrounding the soda lakes Natron (largely in Tanzania) and Magadi (Kenya). Three Alcolapia species are present at Natron (Alcolapia alcalicus, Alcolapia ndalalani and Alcolapia latilabris) and one at Magadi (Alcolapia grahami). All are IUCN Red Listed as either vulnerable or endangered. We performed analyses of morphometric and genetic structure on 13 populations of the Natron Alcolapia flock, and one A. grahami population of Lake Magadi as an out-group. Morphometric analyses revealed significant differentiation in the head and mouth shape of the species at Natron. From a genetic perspective, among 70 mtDNA control region sequences 17 haplotypes were found, showing in the minimum spanning network a star-like pattern around the widespread haplotype 2lat. At Natron, there was limited genetic differentiation between the different populations of A. alcalicus and A. latilabris, despite apparent ecological barriers of extreme alkalinity that suggested their populations were isolated. Instead, there appeared to be some population connectivity, with a rate of 0.5–2.3 migrants per generation suggesting that natural factors, such as intense rains or transmission by large piscivorous birds, facilitate population connectivity and maintain genetic similarity. The outputs of high population connectivity and one genetic unit at the basin level (despite morphological divergence) suggest that any human activities that disrupt the connectivity of the freshwater resources of the Natron catchment could further threaten the integrity and current status of these already threatened fish populations.