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Natural history museums are irreplaceable tools to study and preserve the biological diversity around the globe and among the primary actors in the recognition of species and the logical repositories for their type specimens. In this paper we surveyed the consistency of the preserved specimens of amphibians and reptiles housed in the major Italian scientific collections, and verified the presence of threatened species according to the IUCN Red List, including the Extinct (EX), Extinct in the Wild (EW), Critically Endangered (CR), Endangered (EN), and Vulnerable (VU) categories. Altogether, we analyzed 39 Italian zoological collections. We confirmed the presence of one extinct reptile (Chioninia coctei) and five extinct or extinct in the wild amphibian species (Atelopus longirostris, Nectophrynoides asperginis, Pseudophilautus leucorhinus, P. nasutus, and P. variabilis). Seven CR amphibians, fourteen CR reptile species and the extinct skink C. coctei are shared by more than one institution. Museums which host the highest number of threatened and extinct amphibian species are respectively Turin (17 CR and 1 EX), Florence (13 CR and 1 EX), and Trento (15 CR and 1 EW), while for reptiles the richest museums are those from Genoa (15 CR and 1 EX), Florence (11 CR and 1 EX), and Pisa (7 CR). Finally, we discussed the utility of natural history museums and the strategies to follow for the implementation of their functionality.

We investigated the relationship between age and body length, and age at sexual maturity of Physeter macrocephalus individuals stranded along the Italian coast. Our molecular analysis shows that all our samples belong to the C.001.002 haplotype, shared between Atlantic and Mediterranean populations. We show that males attain sexual maturity at 10 years, similar to those from other marine areas. However, considering the same body length class, Mediterranean males are older than Atlantic ones. Our finding of a Mediterranean pregnant female of only 6.5 m in length and an assessed age of 24–26 years is particularly noteworthy, considering that females reach sexual maturity at about 9 years and 9 m of total length in other regions. Comparing our results with the literature data, we highlight the positive correlation between lifespan, adult body length and weight of males from the Mediterranean and Atlantic Ocean. Regardless of whether the relatively small size of Mediterranean specimens is a consequence of an inbreeding depression or an adaptation to less favorable trophic conditions, we recommend to closely monitor this population from a conservation perspective. In fact, its low genetic diversity likely corresponds to a relatively limited ability to respond to environmental changes compared with other populations.

Cystic Echinococcosis (CE) caused by Echinococcus granulosus sensu lato (s.l.) is one of the most important parasitic zoonotic diseases in the world and it represents an important public health and socio-economic concern. In the Mediterranean basin, CE is widespread and it is endemic in Italy, with major prevalence in southern areas. Several studies have investigated CE in domestic pigs, however, such data in wild boars are scant. In the last decades the wild boar population in Italy has increased and this ungulate could play an important role in the spreading of CE in the wild. Here we report on the prevalence and fertility rate of hydatid cysts in wild boars that were shot during two hunting seasons (2016–2017) in the Campania region of southern Italy.For each animal, a detailed inspection of the carcass and organs (lungs, liver and spleen) was performed and when cysts were found, their number, morphology and fertility were determined by visual and microscopic examination. Cysts were classified morphologically as fertile, sterile, caseous and calcified. Protoscoleces and germinal layers were collected from individual cysts and DNA was extracted to identify different strains/genotypes of E. granulosus s.l.Out of a total of 2108 wild boars 93 (4.4%) were found positive for CE. Infected animals were 45 males and 48 females, aged between 1 and 8 years. The average number of cysts per wild boar was 1.3 (min 1 - max 13). The total number of cysts collected was 123, of which 118 (95.9%) in the liver, 4 (3.3%) in the lungs and 1 (0.8%) in the spleen. Of all analyzed cysts, 70 (56.9%) were fertile and 53 (43.1%) sterile/acephalous. The presence of fertile cysts in 19.4% of CE-positive animals is noteworthy. Overall, molecular diagnosis showed 19 wild boars infected with the pig strain (G7). Keywords: Cystic echinococcosis, Wild boar, Genotyping, Drive hunting, Southern Italy

Background The Mediterranean swordfish stock is overfished and considered not correctly managed. Elucidating the patterns of the Mediterranean swordfish population structure constitutes an essential prerequisite for effective management of this fishery resource. To date, few studies have investigated intra-Mediterranean swordfish population structure, and their conclusions are controversial. Methods A panel of 20 microsatellites DNA was used to investigate fine-scale population structuring of swordfish from six main fishing areas of the Mediterranean Sea. Results This study provides evidence to reject the hypothesis of a single swordfish population within the Mediterranean Sea. DAPC analysis revealed the presence of three genetic clusters and a high level of admixture within the Mediterranean Sea. Genetic structure was supported by significant FST values while mixing was endorsed by the heterozygosity deficit observed in sampling localities indicative of a possible Wahlund effect, by sampling admixture individuals. Overall, our tests reject the hypothesis of a single swordfish population within the Mediterranean Sea. Homing towards the Mediterranean breeding areas may have generated a weak degree of genetic differentiation between populations even at the intra-basin scale.

Abstract Anchovies represent the largest world’s marine fish catches and the current threats on their populations impose a sustainable exploitment based on sound scientific information. In the European anchovy (Engraulis encrasicolus), the existence of several populations has been proposed but a global view is missing. Using a multidisciplinary approach, here we assessed the divergence among different ecotypes and its possible causes. SNPs have revealed two functionally distinct ecotypes overlapping in the Central Mediterranean, with one ecotype confined near the river estuaries. The same SNPs outliers also segregated two distinct populations in the near Atlantic, despite their large spatial distance. In addition, while most studies suggested that adaptation to low salinity is key to divergence, here we show that the offshore ecotype has higher environmental tolerance and an opportunistic feeding behaviour, as assessed by the study of environmental conditions, anchovy diet and trophic levels, and passive egg dispersal. These results provide insights into the anchovy evolutionary history, stressing the importance of behaviour in shaping ecotypes.

Canine vector-borne diseases (CVBD) are an important and emerging health concern for humans and animals worldwide. The purpose of the presented study was to assess, from 2016 to 2019, the seroprevalence of CVBD agents and clarify the epidemiology of tick-borne disease in stray dogs living in the Campania Region, Southern Italy. For this purpose, blood samples were collected from January 2016 to December 2019 from 1023 dogs in authorized kennels located in the five municipalities of the Campania Region. SNAP® 4DX® from IDEXX® Laboratories was used for detection of Ehrlichia spp., Anaplasma spp., Borrelia burgdorferi sensu lato (s.l.), and Dirofilaria immitis antibodies. The overall seroprevalence of CVBD in stray dogs was 19.6% (95% Confidence Intervals (CI): 17.2–22.8%; 201/1023). The most common pathogen was Ehrlichia spp., with a percentage of positivity of 16.03%, followed by Anaplasma spp. with 7.8%. B. burgdorferi s.l. and D. immitis were detected in only 0.2% of dogs; co-infection was detected in 4.5% of stray dogs tested. No link was detected between the gender, age, location, and CVBD seropositivity, except for Ehrlichia spp. for which location (Avellino Province; p = 0.007) and gender (male, p = 0.002) were risk factors for seropositivity. Our results demonstrated that animals are exposed to at least one of the four etiological agents (Ehrlichia spp., Anaplasma spp. Borrelia burgdorferi s.l., and Dirofilaria immitis) transmitted by vectors. Finally, this study highlighted the utility of serological monitoring in stray dogs, housed in kennels, given the threat posed by CVBD to animals and the zoonotic implications of these etiological agents and their vectors on human health.

Skeletal anomalies are commonplace among farmed fish. The pug-headedness anomaly is an osteological condition that results in the deformation of the maxilla, pre-maxilla, and infraorbital bones. Here, we report the first record of pug-headedness in an isolated population of the critically endangered native Mediterranean trout Salmo trutta L., 1758 complex from Sardinia, Italy. Fin clips were collected for the molecular analyses (D-loop, LDH-C1* locus. and 11 microsatellites). A jaw index (JI) was used to classify jaw deformities. Ratios between the values of morphometric measurements of the head and body length were calculated and plotted against values of body length to identify the ratios that best discriminated between malformed and normal trout. Haplotypes belonging to the AD lineage and the genotype LDH-C1*100/100 were observed in all samples, suggesting high genetic integrity of the population. The analysis of 11 microsatellites revealed that observed heterozygosity was similar to the expected one, suggesting the absence of inbreeding or outbreeding depression. The frequency of occurrence of pug-headedness was 12.5% (two out of 16). One specimen had a strongly blunted forehead and an abnormally short upper jaw, while another had a slightly anomaly asymmetrical jaw. Although sample size was limited, variation in environmental factors during larval development seemed to be the most likely factors to trigger the deformities.

The Mediterranean monk seal (Monachus monachus) is one of the most endangered marine mammals in the world. This species has been threatened since ancient times by human activities and currently amounts to approximately 700 individuals distributed in the Eastern Mediterranean Sea (Aegean and Ionian Sea) and Eastern Atlantic Ocean (Cabo Blanco and Madeira). In other areas, where the species is considered “probably extinct”, an increase in sporadic sightings has been recorded during recent years. Sightings and accidental catches of Mediterranean monk seals have become more frequent in the Adriatic Sea, mainly in Croatia but also along the coasts of Montenegro, Albania and Southern Italy. A Mediterranean monk seal pup was recovered on 27 January 2020 on the beach of Torre San Gennaro in Torchiarolo (Brindisi, Apulia, Italy). DNA was extracted from a tissue sample and the hypervariable region I (HVR1) of the mitochondrial DNA control region was amplified and sequenced. The alignment performed with seven previous published haplotypes showed that the individual belongs to the haplotype MM03, common in monk seals inhabiting the Greek islands of the Ionian Sea. This result indicates the Ionian Islands as the most probable geographical origin of the pup, highlighting the need to intensify research and conservation activities on this species even in areas where it seemed to be extinct.

Intense and prolonged mortality caused by over-exploitation could drive the decay of genetic diversity which may lead to decrease species’ resilience to environmental changes, thus increasing their extinction risk. Swordfish is a high commercial value species, especially in the Mediterranean Sea, where it is affected by high catch levels. Mediterranean swordfish consist of a population genetically and biologically distinct from Atlantic ones and therefore managed as a separate stock. The last Mediterranean swordfish stock assessment reported that in the last forty years Mediterranean swordfish has been overfished and, to date, it is still subject to overfishing. A comparison between an available mitochondrial sequence dataset and a homologous current sample was carried out to investigate temporal genetic variation in the Mediterranean swordfish population over near twenty years. Our study provides the first direct measure of reduced genetic diversity for Mediterranean swordfish during a short period, as measured both in the direct loss of mitochondrial haplotypes and reduction in haplotype diversity. A reduction of the relative females’ effective population size in the recent sample has been also detected. The possible relationship between fishery activities and the loss of genetic diversity in the Mediterranean swordfish population is discussed.

The Mediterranean brown trout (Salmo trutta complex) is one of the freshwater fish species complex at greater risk of extinction in the Mediterranean area. The introduction of alien invasive species and their interaction with the native fauna represent some of the major threats to the survival of this species. Currently, the genetic variability of the Mediterranean trout (Salmo trutta complex) is being compromised by the introgressive hybridization with the Atlantic trout (Salmo trutta Linnaeus, 1758). Therefore, it is necessary to gain further knowledge on genetic and demographic characteristics of Mediterranean trout populations, that, combined with environmental and angling data, will allow to undertake sound conservation strategies. The aims of the present study were to analyze: i) the demographic characteristics of the Mediterranean trout in seven central Apennine river basins where native populations are threatened by stocking with the domestic trout of north Atlantic origin; ii) the influence of both environmental parameters and different fishery management strategies on the status of native trout populations. The project focuses on 14 sites included in the Natura 2000 EU wide network of nature protection areas, established under the 1992 Habitat Directive. A total of 25 watercourses were investigated for a total of 32 sampling sites. A census of the fish fauna was carried out by electrofishing at each sampling location. Fish and environmental data were collected in spring and autumn 2014. The results obtained in the present research allowed us to detect the presence of three residual Mediterranean trout populations with a high degree of genetic integrity in the study area. The native trout populations were characterized by higher abundances and higher adult density values. Well-structured native populations with higher legal-size specimens density and poor body conditions in terms of relative weight were observed in the no-fishing areas, probably due to the presence of intraspecific competition phenomena. The environmental analysis confirmed the key role played by the geological characteristics of the central Apennines in shaping the introgression levels observed in the wild trout populations, while the recovery of the water quality appears decisive in the development of management strategies aimed at their conservation.

In the present study the genetic variability of European sardine from Adriatic and Ionian seas was investigated in order to detect the occurrence of genetic structure within and between these basins. In several samples the analysis of genetic variability at eight microsatellite loci showed a number of homozygote individuals higher than expected at Hardy-Weinberg equilibrium. The inter-population differentiation level estimated by AMOVA, qST and rRST and Bayesian descriptors detected no signs of population differentiation between the samples analysed. These results are consistent with previous studies based on allozymes and several mitochondrial DNA markers and add further evidence contradicting the early identification, based on morphological and reproductive data, of two sub-populations in the Adriatic Sea.

It is well known that temporal fluctuations in small populations deeply influence evolutionary potential. Less well known is whether fluctuations can influence the evolutionary potentials of species with large census sizes. Here, we estimated genetic population parameters from as survey of polymorphic microsatellite DNA loci in archived otoliths from Adriatic European anchovy (Engraulis encrasicolus), a fish with large census sizes that supports numerous local fisheries. Stocks have fluctuated greatly over the past few decades, and the Adriatic fishery collapsed in 1987. Our results show a significant reduction of mean genetic parameters as a consequence of the population collapse. In addition, estimates of effective population size (Ne) are much smaller than those expected in a fishes with large population census sizes (Nc). Estimates of Ne indicate low effective population sizes, even before the population collapse. The ratio Ne/Ne ranged between 10-6 and 10-8, indicating a large discrepancy between the anchovy gene pool and population census size. Therefore, anchovy populations may be more vulnerable to fishery effort and environmental change than previously thought.

In this pilot study for the first time, ancient DNA has been extracted from bone remains of Salmo trutta. These samples were from a stratigraphic succession located in a coastal cave of Calabria (southern Italy) inhabited by humans from upper Palaeolithic to historical times. Seven pairs of primers were used to PCR-amplify and sequence from 128 to 410 bp of the mtDNA control region of eleven samples. Three haplotypes were observed: two (ADcs-1 and MEcs-1) already described in rivers from the Italian peninsula; one (ATcs-33) belonging to the southern Atlantic clade of the AT Salmo trutta mtDNA lineage (sensu Bernatchez). The prehistoric occurrence of this latter haplotype in the water courses of the Italian peninsula has been detected for the first time in this study. Finally, we observed a correspondence between frequency of trout remains and variation in haplotype diversity that we related with ecological and demographic changes resulting from a period of rapid cooling known as the Younger Dryas.

The sustained exploitation of marine populations requires an understanding of a species' adaptive seascape so that populations can track environmental changes from short- and long-term climate cycles and from human development. The analysis of the distributions of genetic markers among populations, together with correlates of life-history and environmental variability, can provide insights into the extent of adaptive variation. Here, we examined genetic variability among populations of mature European anchovies (n = 531) in the Adriatic (13 samples) and Tyrrhenian seas (2 samples) with neutral and putative non-neutral microsatellite loci. These genetic markers failed to confirm the occurrence of two anchovy species in the Adriatic Sea, as previously postulated. However, we found fine-scale population structure in the Adriatic, especially in northern areas, that was associated with four of the 13 environmental variables tested. Geographic gradients in sea temperature, salinity and dissolved oxygen appear to drive adaptive differences in spawning time and early larval development among populations. Resolving adaptive seascapes in Adriatic anchovies provides a means to understand mechanisms underpinning local adaptation and a basis for optimizing exploitation strategies for sustainable harvests.

The Mediterranean brown trout (Salmo trutta complex) is one of the freshwater fish species complex at greater risk of extinction in the Mediterranean area. The introduction of alien invasive species and their interaction with the native fauna represent some of the major threats to the survival of this species (Caputo et al., 2004). Currently, the genetic integrity of the Mediterranean trout is being compromised by the introgressive hybridization with the Atlantic trout. This hybridization, which was favored by stocking programs carried out in the last century (Splendiani et al., 2013), is resulting in the loss of native genetic variability. The Life+ TROTA Project, which started in November 2013, is aimed to preserve the last populations of Apennine trout (Salmo cettii Rafinesque, 1810) and recover autochthonous genetic diversity. These action are being carried out in seven river basins of central Italy, where native populations have been threatened by stocking with Atlantic trout. The data collected are the indispensable premise for the adoption of the necessary strategies for conservation of the Apennine trout. The trout is a wide-ranging species complex native to Eurasia and North Africa. The northern limits to its range are represented by Iceland, northern Scandinavia and Russia, while the southern limits are represented by the mountain streams of northern Morocco. This species has also been introduced in several other locations around the world (North and South - America, Australia, etc). The Apennine trout is considered as critically endangered in the IUCN Red List of Italian vertebrates (Rondinini et al., 2013) due to water abstraction, overfishing and stocking with non-native trout (resulting in hybridization and competition). In the last 10 - 12 years the population has declined by almost 30%, and this trend is expected also in the future as water abstraction will most likely increase. The project focuses on 14 sites included in the Natura 2000 EU wide network of nature protection areas, established under the 1992 Habitat Directive. The project involved also the territory of the National Park of Monti Sibillini. The study area comprises 7 watersheds: Metauro, Cesano, Esino, Potenza, Chienti, Tevere and Tenna; a total of 25 watercourses were investigated with 32 sampling stations. A census of the fish fauna by electrofishing was carried out at each sampling location. For each specimen captured, total length and weight was measured, and a fin clip was collected for genomic DNA extraction. Genetic analyses were performed by using mitochondrial (D-loop fragment of 310 bp) and nuclear markers (locus LDH-C1* and 12 microsatellites). Bayesian clustering implemented in Structure 2.3.2.1 was used for estimating the number of groups (k) represented by all sampled individuals (Pritchard et al., 2000; Falush et al., 2003). The genetic and demographic characterization of trout populations in 32 sites allowed us to select: i) six sites for the collections of adults, needed for the production of genetically pure juveniles trout, ii) four sites for supportive breeding actions and iii) six sites to carry out the eradication of alien trout and successive re-introduction of Mediterranean trout. Another significant activity of the project is the hydrological characterization of the water courses by the Minimum Flow Analysis (MFA), adopting the MesoHABSIM method (Parasiewicz, 2007). This action is crucial to monitor the effect of water abstraction in the sites involved in the concrete conservation actions (e.g., eradication and re-introduction), and more generally, in the study area. In this way, we hope to increase the chances of long term survival of Apennine trout populations. Main expected results are the following:  Identification of natural populations of Apennine trout in the study area and their protection through specific changes in existing regulation.  Establishment of genetically pure wild populations with fish produced in an hatchery specifically adapted to manage wild native broodstock.  Widening Apennine trout habitat availability by removal of non native trout in areas to be restocked.  Restocking of streams selected in previous actions with pure juvenile trout.  Assessment of the summer river flows in order to increase the chances of survival of Apennine trout populations in compliance with the regulations on minimum flow and water quality.  Involvement of local sport fishing community to increase their awareness of the problem and ease their acceptance of more restricting fishery and fish management rules. At present the genetic and demographic characterization of the trout populations is completed. Table 1 and Figures 1-2 show the results of abundance analysis of the fish assemblages divided by basin. The standing crop’s mean values can be considered in the standard range, according to the small size and the limited productivity that generally characterized the watercourses analyzed, as most of the Apennine rivers. The differences of the mean values among basins were highly statistically significant according to the ANOVA analysis (density: F= 5.24, p=0.001; standing crop: F=12.42, p=0.001). The results of genetic analysis separated clearly native and aliene genomes (K = 2, Fig. 3a) and showed the presence of three distinct genetic stocks of native Apennine trout (K = 4, Fig. 3b): i) Tevere (green bars), ii) Tenna (red) and iii) Chienti /Potenza/Metauro/Esino (yellow). Populations characterized by very low introgression values will be the source of wild individuals to produce pure juvenile trouts in captivity. Four moderately introgressed populations will be subject to supportive breeding activities. Finally, six stream, where trouts showed almost exclusively an alien genetic make-up will be selected for the eradication activities. The data collected are the indispensable premise for the adoption of the necessary strategies for conservation of the Apennine trout in Central Italy.

Parthenogenesis, unisexuality and triploidy are interesting but poorly studied phenomena occurring in some reptile species. The mourning gecko (Lepidodactylus lugubris) represents a complex of diploid and triploid parthenogenetic mostly all-female populations (males occur quite rarely) widely distributed in coastal areas of the Indian and Pacific Oceans. Here, we study karyotypes of a male and two female L. lugubris (LLU) triploid individuals (3n = 66) using comparative painting with Gekko japonicus, Hemidactylus turcicus and H. platyurus chromosome specific probes to visualize the homologous regions and to reveal genus specific rearrangements. Also, we applied a 28S ribosomal DNA probe and Ag-staining to detect nucleolus organizer regions (NORs). Our results suggest that the karyotype of L. lugubris underwent a chromosome fission and a fusion after its divergence from a common ancestor of the Gekko-Hemidactylus group. The NORs were found to be located on one out of three homologs on each of LLU8, LLU15 and LLU18, thus further confirming a hybrid origin of triploid individuals. It seems that three different bisexual populations might have contributed to the origin of this triploid parthenogenetic population. We postulate that the heterozygosity in NOR localization is maintained in the triploid clone studied by the absence of recombination as described in whiptail lizards. The pattern of NOR localizations and homologous regions in males and females, as well as the absence of other detectable karyotypic differences, suggest that males arise spontaneously in all female populations and do not arise from independent hybridizations with different species.

Caratterizzazione aplotipica di una balenottera comune spiaggiata presso Ancona (Mare Adriatico, Italia centrale). Riassunto La sequenza della regione di controllo del DNA mitocondriale di un esemplare di balenottera comune (Balaenoptera physalus) spiaggiata presso Ancona (Mare Adriatico) viene descritta e confrontata con quelle note per la specie. L’aplotipo ottenuto è risultato identico a quello descritto per il Mar Ligure, permettendo di ipotizzare spostamenti fra l’area tirrenica e l’Adriatico centro-settentrionale.

A study on chromosomes of two Scincus populations attributed to S. scincus and S. albifasciatus was carried out by conventional Giemsa staining. Both samples showed the same diploid number of 2N = 32 and identical chromosome morphology. Therefore, these populations seem to be lacking in chromosome changes that could act as barriers to gene flow. However, because speciation in lizards is not always associated with chromosome repatterning, the taxonomic status of “scincus” and “albifasciatus” forms remains unsettled.

Natural history museums are irreplaceable tools to study and preserve the biological diversity around the globe and among the primary actors in the recognition of species and the logical repositories for their type specimens. In this paper we surveyed the consistency of the preserved specimens of amphibians and reptiles housed in the major Italian scientific collections, and verified the presence of threatened species according to the IUCN Red List, including the Extinct (EX), Extinct in the Wild (EW), Critically Endangered (CR), Endangered (EN), and Vulnerable (VU) categories. Altogether, we analyzed 39 Italian zoological collections. We confirmed the presence of one extinct reptile (Chioninia coctei) and five extinct or extinct in the wild amphibian species (Atelopus longirostris, Nectophrynoides asperginis, Pseudophilautus leucorhinus, P. nasutus, and P. variabilis). Seven CR amphibians, fourteen CR reptile species and the extinct skink C. coctei are shared by more than one institution. Museums which host the highest number of threatened and extinct amphibian species are respectively Turin (17 CR and 1 EX), Florence (13 CR and 1 EX), and Trento (15 CR and 1 EW), while for reptiles the richest museums are those from Genoa (15 CR and 1 EX), Florence (11 CR and 1 EX), and Pisa (7 CR). Finally, we discussed the utility of natural history museums and the strategies to follow for the implementation of their functionality. [ABSTRACT FROM AUTHOR]