Vulgaris Meridionalis Research Articles

Fine-scale spatial genetic structure and dispersal among Italian smooth newt populations in a rural landscape

Amphibians are particularly sensitive to habitat loss and fragmentation caused by the intensification and modernization of farming occurring in the second half of the twentieth century in the Mediterranean basin. However, artificial water bodies, associated with traditional husbandry, proved to be important surrogate for amphibian feeding and reproduction. Here, multilocus genotypes were used to investigate the spatial population structure of Lissotriton vulgaris meridionalis and the role of drinking troughs in supporting viable breeding populations within a rural landscape interested by traditional husbandry and agriculture. Our genetic analysis highlighted the conservation value and the potential stepping-stone function of artificial aquatic sites in the dispersal of the species and for the gene flow maintenance. Indeed, populations of drinking troughs show allelic richness and heterozygosity levels comparable to those from natural ponds and there is no great evidence of genetic bottlenecks. A complex system of artificial aquatic sites and few natural wetlands was identified sustaining a well-structured network of demes highly interconnected with themselves and natural aquatic sites. The conservation of the identified genetic clusters may be useful to prevent further population declines and future loss of genetic diversity within the study area characterized by scarce natural wetlands that frequently dried because of agricultural practices and strong seasonality. Site-specific protection measures are needed to contrast the progressive disappearance of drinking troughs observed in the last years in Italy because of the abandonment of traditional farming practices in favour of modern agriculture and intensive farming.

Monitoring newt communities in urban area using eDNA metabarcoding.

Newts are amphibians commonly present in small ponds or garden pools in urban areas. They are protected in many countries and their presence is monitored through visual observation and/or trapping. However, newts are not easy to spot as they are small, elusive and often hidden at the bottom of water bodies. In recent years, environmental DNA (eDNA) has become a popular tool for detecting newts, with a focus on individual species using qPCR assays. Here, we assess the effectiveness of eDNA metabarcoding compared to conventional visual surveys of newt diversity in 45 ponds within urban areas of Geneva canton, Switzerland. We designed newt-specific mitochondrial 16S rRNA primers, which assign the majority of amplicons to newts, and were able to detect four species known to be present in the region, including the invasive subspecies Lissotriton vulgaris meridionalis, native to the Italian peninsula, that has been introduced in the Geneva area recently. The obtained eDNA results were congruent overall with conventional surveys, confirming the morphological observations in the majority of cases (67%). In 25% of cases, a species was only detected genetically, while in 8% of cases, the observations were not supported by eDNA metabarcoding. Our study confirms the usefulness of eDNA metabarcoding as a tool for the effective and non-invasive monitoring of newt community and suggests its broader use for the survey of newt diversity in urban area at larger scales.

Can We Share? Feeding Strategy in Three Syntopic Newts in Artificial Habitats

Natural aquatic sites are disappearing worldwide, especially in the Mediterranean region where amphibians are frequently forced to move for reproduction to artificial sites designed for irrigation and cattle watering (i.e., wells, tanks and drinking troughs). In artificial aquatic sites, where resources (space and food) are usually limited, trophic niche information can be particularly useful to infer the suitability of habitats for amphibian conservation especially when more than one species co-occurs. In this paper, we focused on three newt species: The Italian newt (Lissotriton italicus), the Italian smooth newt (Lissotriton vulgaris meridionalis) and the Italian crested newt (Triturus carnifex) inhabiting man-made wells widespread in an area in Central Italy characterized by few available natural aquatic sites. Specifically, we analyzed the trophic spectrum of the species, their interactions and overlap, and discussed the potential role of wells in amphibian conservation. Overall, 550 newt individuals occurring in 16 distinct wells were sampled. The study species consumed similar resources, mainly of aquatic origin, with Diptera larvae and Cladocera representing the most important preys. The high degree of diet overlap observed may be due to site oligotrophy and high availability of small-sized prey, and it does not necessarily lead to competition. Newts had similar narrow niche width values and a generalist feeding pattern with high diversity among individuals. Lissotriton italicus and T. carnifex showed wider niche width in isolation than in syntopy condition, probably as a result of interspecific competition and/or intraguild predation. We showed that artificial aquatic sites are important for newt ecology and conservation since they allow up to three species to cohabit, thus representing a good surrogate of natural habitats. The study wells apparently provided suitable trophic conditions for newts in terms of prey availability and catchability. To date, just a few studies have contributed to a greater understanding of newts’ diet in artificial aquatic sites and this gap of knowledge has to be filled to clarify their role in amphibian ecology and conservation.

Can a cow save a newt? The role of cattle drinking troughs in amphibian conservation

Abstract Amphibians are the vertebrate group experiencing the steepest global population decline and species loss. Habitat alteration and loss caused by the intensification of agriculture is among the main causes; however, in the past, amphibians have been favoured by traditional agriculture and husbandry as more terrestrial and aquatic habitats became available through land‐use diversification and the construction of artificial facilities for cattle watering. Troughs for cattle watering may represent the only permanent water points for amphibian reproduction in human‐modified landscapes. The present study focused on Lissotriton vulgaris meridionalis, a semi‐aquatic salamander, subendemic and widespread in Italy, capable of colonizing artificial aquatic habitats. The main goal was to determine the importance of drinking troughs for the ecology and conservation of the species in a hilly area (Latium region, Central Italy) subjected to traditional livestock farming (i.e. Maremmana cattle). Specifically, generalized linear models (GLMs) were used to analyse the effects of drinking trough characteristics (i.e. size, substrate, aquatic vegetation), landscape features (distance to the forest, forest cover, percentage of cultivated lands and infrastructural development, terrestrial connectivity) and intensity of management practices on the occurrence and abundance of the target species. In the light of the present results, it is suggested that drinking troughs represent suitable surrogates for pristine habitats for L. vulgaris meridionalis. Hence, even man‐made aquatic habitats, associated with traditional husbandry, may help to sustain amphibian populations in landscapes where natural wetlands are scarce. The present findings are timely because of the lack of empirical data on the role of drinking troughs in amphibian conservation within rural socio‐economic scenarios, which may have been neglected in research and policy in favour of lakes, rivers and streams.

Herps without borders: a new newt case and a review of transalpine alien introductions in western Europe

Abstract Biogeographic processes have led to different evolutionary taxa occurring in the northern and southern edges of the Alpine Mountains in Western Europe. The integrity of this diversity is being challenged by frequent human-mediated trans-alpine translocations, sometimes leading to biological invasions. Several alien terrestrial vertebrates of south Alpine origins (Italy, Swiss Ticino) are causing damages to native north Alpine fauna. In this paper, we used molecular tools to characterize the understudied case of the Mediterranean smooth newt (Lissotriton vulgaris meridionalis) expanding in the outskirts of Geneva since its introduction before 1975. Mitochondrial and nuclear DNA sequencing suggest that these exotic populations are a mixture between two diverged L. v. meridionalis lineages from central Italy, and traces of potential hybridization with the native L. v. vulgaris was detected. This situation echoes many other trans-alpine alien introductions. We review all comparable cases of southern to northern Alps introductions in vertebrates, including seven reptiles and four amphibians. The majority of south alpine alien lineages were presumably imported voluntarily by enthusiasts and appear to perform better in the disturbed habitats found in the anthropogenic landscapes of Western Europe compared to their native north Alpine counterparts. Most pose serious threats to related species of similar ecology, through direct competition, predation and introgressive hybridization. Difficulties to detect alien species on time lead to significant conservation costs. Better education together with more appropriate and reactive management plans will be necessary to limit the impact of future alien introductions.

An improved microsatellite panel to assess genetic variability of the Italian smooth newt (Lissotriton vulgaris meridionalis)

Habitat loss and fragmentation are threatening amphibians by increasing population isolation. However, artificial waterbodies created for livestockmay contrast this phenomenon by providing surrogate habitats for amphibians. Here, we performed a genetic study on an amphibian species, Lissotriton vulgaris meridionalis, in a rural area in central Italy where natural wetlands are disappearing and drinking troughs for cattle watering are widespread. Specifically, we tested a panel of microsatellite markers to identify a suitable tool for addressing conservation genetic issues of this species that is undergoing severe local decline. Twelve of the 20 tested loci produced reliable amplifications and were polymorphic. Three distinct units with a low level of gene flow were distinguished and the population genetic structuring overlapped with geographic distribution. Such loci will be useful to assess the genetic diversity of the species across multiscale levels for its management and conservation.

Unraveling climate influences on the distribution of the parapatric newts Lissotriton vulgaris meridionalis and L. italicus

BackgroundClimate is often considered as a key ecological factor limiting the capability of expansion of most species and the extent of suitable habitats. In this contribution, we implement Species Distribution Models (SDMs) to study two parapatric amphibians, Lissotriton vulgaris meridionalis and L. italicus, investigating if and how climate has influenced their present and past (Last Glacial Maximum and Holocene) distributions. A database of 901 GPS presence records was generated for the two newts. SDMs were built through Boosted Regression Trees and Maxent, using the Worldclim bioclimatic variables as predictors.ResultsPrecipitation-linked variables and the temperature annual range strongly influence the current occurrence patterns of the two Lissotriton species analyzed. The two newts show opposite responses to the most contributing variables, such as BIO7 (temperature annual range), BIO12 (annual precipitation), BIO17 (precipitation of the driest quarter) and BIO19 (precipitation of the coldest quarter). The hypothesis of climate influencing the distributions of these species is also supported by the fact that the co-occurrences within the sympatric area fall in localities characterized by intermediate values of these predictors. Projections to the Last Glacial Maximum and Holocene scenarios provided a coherent representation of climate influences on the past distributions of the target species. Computation of pairwise variables interactions and the discriminant analysis allowed a deeper interpretation of SDMs’ outputs. Further, we propose a multivariate environmental dissimilarity index (MEDI), derived through a transformation of the multivariate environmental similarity surface (MESS), to deal with extrapolation-linked uncertainties in model projections to past climate. Finally, the niche equivalency and niche similarity tests confirmed the link between SDMs outputs and actual differences in the ecological niches of the two species.ConclusionsThe different responses of the two species to climatic factors have significantly contributed to shape their current distribution, through contractions, expansions and shifts over time, allowing to maintain two wide allopatric areas with an area of sympatry in Central Italy. Moreover, our SDMs hindcasting shows many concordances with previous phylogeographic studies carried out on the same species, thus corroborating the scenarios of potential distribution during the Last Glacial Maximum and the Holocene emerging from the models obtained.

Northern richness and cryptic refugia: phylogeography of the Italian smooth newtLissotriton vulgaris meridionalis

Recent phylogeographical studies have re-evaluated the role of refugia in central and northern Europe for glacial persistence and postglacial assembly of temperate biota. Yet, on a regional scale within Mediterranean peninsulas, putative ‘northern’ refugia's contribution to the current structure of biodiversity still needs to be fully appreciated. To this end, we investigated the phylogeographical structure and the evolutionary history of the Italian smooth newt, Lissotriton vulgaris meridionalis, through phylogeographical, molecular dating and historical demographic analyses. We found ten differentiated mitochondrial lineages with a clear geographical association, mainly distributed in northern Italy. The most ancient divergence among these lineages was estimated at the Early Pleistocene and was followed by a series of splits throughout the Middle Pleistocene. No haplogroup turned out to be derived from another one, each one occupying terminal positions within the phylogenetic network topologies. These results suggest an unprecedented scenario involving long-term survival of distinct evolutionary lineages in multiple northern Mediterranean refugia. This scenario mirrors on a smaller geographical scale what has been previously observed in the literature concerning northern European environments; it also sheds more light on how northern Italy has contributed to temperate species' long-term survival and to the assembly of regional biota. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 113, 590–603.

Scale‐dependent environmental variables affecting red squirrel (Sciurus vulgaris meridionalis) distribution

We investigated the effects of habitat fragmentation on the endemic subspecies of red squirrel Sciurus vulgaris meridionalis in the Pollino National Park, Calabria, Southern Italy. Presence/absence of squirrels was monitored using drey (nest) counts in 51 1‐ha census plots. Squirrel dreys were found in 16 plots (31%). Patch size was not correlated to squirrel presence. Squirrels were found in patches ranging from 3.19 to 6051 ha. Small‐scale forest structure significantly affected the probability of occurrence. The proportion of conifers and average tree height positively predict squirrel presence; furthermore, nest density was positively correlated with high tree species diversity and the proportion of deciduous oaks (Quercus cerris, Q. ilex). Also at the home‐range scale the proportion of conifer forest and oak‐dominated deciduous forests positively predicted squirrel presence (200–300 m radius). At the even larger scale, corresponding with potential dispersal distances (3000 m radius), landscape parameters indicating a lower degree of fragmentation and proportion of oak seemed to favour squirrel presence. Our results emphasize that multi‐scale analyses can enhance our understanding of red squirrel distribution, and that their distribution and abundance were mainly determined by forest structure components, such as food availability, at the home‐range scale. We underline the importance of protection, and eventually increasing conifer and deciduous oak woods range in the Pollino National Park for the management and conservation of endemic Calabrian red squirrels.

Sexual size and shape evolution in European newts (Amphibia: Caudata: Salamandridae) on the Balkan Peninsula

We used a phylogenetic perspective in an examination of the direction and extent of sexual dimorphism in body size and body shape in European newts from the Balkan Peninsula (alpine newts, Mesotriton alpestris; crested newts, Triturus cristatus superspecies; smooth newts, Lissotriton vulgaris). We found a strong, female-biased sexual size dimorphism (SSD) in the analysed clades of alpine newt, whereas within crested newts we found a less stringent female-biased SSD in Triturus carnifex, Triturus macedonicus and Triturus karelinii, and no significant SSD in T. cristatus or Triturus dobrogicus. Among the smooth newts, we found male-biased SSD in Lissotriton vulgaris vularis and Lissotriton vulgaris greacus and no SSD in Lissotriton vulgaris meridionalis. Most of these newts also exhibit a significant sexual dimorphism in body shape, which varied more randomly than body size, regardless of SSD level. Female and male body size as well as the degree of SSD displayed statistically significant phylogenetic signal, while sexual dimorphism in body shape was phylogenetically independent. The relationship between independent contrast data for female size and male size indicated that SSD in European newts could be driven by a disproportionate increase in female size as increase in female size was not accompanied by a proportional increase in male size.

Sexual Dimorphism inTriturus Vulgaris Merindionalisfrom Northern Istria (Slovenia)

Meristic characteristics of Triturus vulgaris meridionalis were measured for 48 males and 38 females from the Mediterranean part of Slovenia. T. v. meridionalis is highly dimorphic in all measured characters, probably as the result of different selective forces acting on each sex. The mean snout-vent lengths were significantly larger in females, also females had wider heads and longer trunks. Males had longer heads and tails.

Habitat use and population density of the red squirrel, Sciurus vulgaris meridionalis, in the Sila Grande mountain range (Calabria, South Italy)

Counts of squirrel nests (dreys) and vegetation surveys in 32 1‐ha census stations were carried out to estimate densities in different forest types and determine habitat selection of the subspecies of red squirrel Sciurus vulgaris meridionalis in the Sila Grande forests, Calabria, South Italy. Dreys were not built randomly, but the majority was constructed at 60–70% of the total height of the nest‐tree, pines and oaks being the preferred tree species. Mean drey densities, 2.75/ha, 2.78/ha, and 2.00/ha in, respectively, Cal‐abrian black pine forest, beech dominated mixed forest, and chestnut woodlands, did not differ between forest types and extrapolated to estimated squirrel densities ranging from 0.44 to 0.61 squirrels/ha. Variation between stations in a tree species diversity index and in distance from the nearest plot of pine woodland significantly affected variation in drey density: squirrels preferred forest habitats dominated by beech, with also black pine and oak available, or chestnut woodlands that were within the range of one (male) squirrel home range diameter (310 m) of a plot of pine forest (mean drey density 5.4/ha). Fewer dreys were found in deciduous (beech or chestnut) woodland >310 m away from areas with pine forest (1.1 dreys/ha). Hence, the availability, at least in the vicinity of the census station, of pine seeds of the Calabrian black pine as main food resource strongly influenced habitat choice.

Body size, age and fecundity variation in different populations of the smooth newt Triturus vulgaris meridionalis in central Italy

Adult and larval body size, age distribution and female fecundity, traits related to fitness in the smooth newt Triturus vulgaris meridionalis, were compared in populations differing in water stability conditions in the area surrounding Rome, near the southern limit of the distribution of this subspecies. Larval body size varied among populations at the end of metamorphosis and was positively related to water stability conditions, which probably led to differences in adult body size. In populations of temporary ponds, age at first breeding was lowered to two years. Traits of female fecundity correlated positively with body size, but not with age. The Italian smooth newt adapted to the unpredictable environment by optimizing reproductive strategies such as the exploitation of the short wet period by means of early deposition of large eggs, which can produce large larvae at metamorphosis.

Analysis of amphibian habitat preferences in a farmland area (Po plain, northern Italy)

AbstractForty-two water bodies in an agricultural lowland area of northern Italy were investigated with respect to the presence of amphibians and to a number of habitat features. The following species were found: Rana kl. esculenta, R. latastei, Bufo bufo, B. viridis, Hyla arborea, Triturus carnifex and T. vulgaris meridionalis. A discriminant analysis was carried out for each species to test whether there were significant difference between ecological characteristics of used and unused sites; all analyses showed a significant difference, and they also made it possible to identify the most important habitat features for each species found. Vegetational factors seemed to be the most important.

A newt ribozyme: a catalytic activity in search of a function.

We analyzed the cleavage properties and the transcription regulation of the newt (Triturus vulgaris meridionalis) self-cleaving RNA. In vitro self-cleavage of model oligoribonucleotides occurs within a double hammerhead structure. In addition, an entire ribozyme molecule, as well as its catalytic domain, "trans-cleaves" in vitro appropriate oligoribonucleotide substrates. Signals encoded within the ribozyme DNA sequences regulate the ribozyme transcription, which is RNA polymerase II dependent. Finally, the deduced secondary structure of the self-cleaving RNA appears to be conserved in evolutionarily distant newt species. These features suggest that the newt ribozyme could play some role in the cell, possibly related to its cleavage properties.

Tooth morphology inTriturus vulgaris meridionalis(Amphibia Urodela) during larval development and metamorphosis

Abstract Arrangement and morphology of teeth in larval and metamorphic Triturus vulgaris meridionalis were studied at the scanning electron microscope and compared with adult and neothenic newts. Monocuspid larval teeth are monostichous on margin of jaws and polystichous on palatine and coronoid bone. They are substituted during metamorphosis by bicuspid teeth arranged in a single row on margin of the jaws and in a double row on the vomerine bones. Bicuspid teeth have initially the same shape but later are dimorphic, according to their distribution in the mouth. The variation of tooth morphology is discussed and correlated with feeding strategies.

Two dispersed highly repeated DNA families of Triturus vulgaris meridionalis (Amphibia, Urodela) are widely conserved among Salamandridae

Two BamHI families of repeated sequences were characterized from the genome of the Italian smooth newt, Triturus vulgaris meridionalis (Amphibia, Urodela). The first family, which is divided into subfamilies, consists of tandemly arranged arrays whose basic repeat is around 398 bp long; these arrays are dispersed throughout the entire chromosome sets of the various species of Triturus tested. Moreover the family is widely conserved among Salamandridae, being detected by genomic DNA blotting of Notophthalmus viridescens, Taricha granulosa, Salamandrina terdigitata and Euproctus platycephalus. The second BamHI family is represented by a cloned sequence of 419 bp, which is dispersed in the chromosome set of several species of Triturus. The sequence is also conserved in S. terdigitata and in E. platycephalus but is not detectable in N. viridescens or T. granulosa. The cloned sequence is most probably only part of a longer unit interspersed within the Triturus genome.

Extra-ribosomal spacer sequences in Triturus

We show that, in Triturus vulgaris meridionalis, sequences homologous to the rDNA “nontranscribed” spacer (NTS) are clustered at chromosomal loci where they are not associated with 18 S or 28 S rDNA genes: these sequences are referred to as the extra-ribosomal spacer sequences. Genomic clones containing such extra-ribosomal spacer sequences have been isolated. As shown by restriction mapping, these clones appear to consist mostly of repetitive BamHI fragments that are, in turn, internally repetitious and highly homologous to each other. The structure of the clones was confirmed by nucleotide sequence analysis, which also demonstrates the high degree of conservation between the BamHI elements and the homologous NTS sequences. An intriguing 12 base-pair homology between the extra-ribosomal spacer sequences and a Xenopus NTS enhancer sequence is reported. The possibility that a repetitive octanucleotide motif found within the BamHI elements could act as a recombination hotspot by virtue of its similarity with the Escherichia coli χ sequence is discussed.

Heterochromatic DNA in Triturus (Amphibia, Urodela) II. A centromeric satellite DNA.

The MspI family of highly repeated sequences is a centromeric satellite DNA representing about 1% of the genome of the Italian smooth newt, Triturus vulgaris meridionalis. We have studied the structure, genomic organization, chromosomal localization and conservation across species of this family. MspI sequences are around 197 bp long, as shown by sequencing of three cloned units. The family is organized in large clusters of tandemly arrayed units, present at almost all the centromeres of T.v. meridionalis, and is well conserved in the T.v. vulgaris subspecies. Conserved MspI sequences are also present in the related species T. helveticus, where they appear to be clustered at the centromeres of only a few chromosomes. MspI sequences are not found in other Triturus species analysed. The correlation of these sequences with the overall distribution pattern of heterochromatin and the extent of their conservation within the genus Triturus, are discussed.

Genomic organization of ribosomal DNA and related sequences inTriturus

The ribosomal RNA genes ofTriturus vulgaris meridionalis are clustered at variable and often multiple chromosomal loci. The rDNA repeats exhibit only a limited and discrete length heterogeneity which is accounted for by the non-transcribed spacer (NTS). Interestingly, sequences homologous to the NTS are clustered outside the ribosomal loci. Clones containing such 'non ribosomal sequences' have been isolated from a genomic library ofT. v. meridionalis and analyzed by restriction mapping. These sequences appear to consist mostly of repetitive Bam HI fragments ranging from 500 bp to 1000 bp. The Bam HI fragments are internally repetitious and highly homologous to each other.