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.

Microsatellite Characterization and Panel Selection for Brown Bear (Ursus arctos) Population Assessment.

An assessment of the genetic diversity and structure of a population is essential for designing recovery plans for threatened species. Italy hosts two brown bear populations, Ursus arctos marsicanus (Uam), endemic to the Apennines of central Italy, and Ursus arctos arctos (Uaa), in the Italian Alps. Both populations are endangered and occasionally involved in human-wildlife conflict; thus, detailed management plans have been in place for several decades, including genetic monitoring. Here, we propose a simple cost-effective microsatellite-based protocol for the management of populations with low genetic variation. We sampled 22 Uam and 22 Uaa individuals and analyzed a total of 32 microsatellite loci in order to evaluate their applicability in individual identification. Based on genetic variability estimates, we compared data from four different STR marker sets, to evaluate the optimal settings in long-term monitoring projects. Allelic richness and gene diversity were the highest for the Uaa population, whereas depleted genetic variability was noted for the Uam population, which should be regarded as a conservation priority. Our results identified the most effective STR sets for the estimation of genetic diversity and individual discrimination in Uam (9 loci, PIC 0.45; PID 2.0 × 10-5), and Uaa (12 loci, PIC 0.64; PID 6.9 × 10-11) populations, which can easily be utilized by smaller laboratories to support local governments in regular population monitoring. The method we proposed to select the most variable markers could be adopted for the genetic characterization of other small and isolated populations.

First core microsatellite panel identification in Apennine brown bears (Ursus arctos marsicanus): a collaborative approach.

BACKGROUND: The low cost and rapidity of microsatellite analysis have led to the development of several markers for many species. Because in non-invasive genetics it is recommended to genotype individuals using few loci, generally a subset of markers is selected. The choice of different marker panels by different research groups studying the same population can cause problems and bias in data analysis. A priority issue in conservation genetics is the comparability of data produced by different labs with different methods. Here, we compared data from previous and ongoing studies to identify a panel of microsatellite loci efficient for the long-term monitoring of Apennine brown bears (Ursus arctos marsicanus), aiming at reducing genotyping uncertainty and allowing reliable individual identifications overtimes. RESULTS: We examined all microsatellite markers used up to now and identified 19 candidate loci. We evaluated the efficacy of 13 of the most commonly used loci analyzing 194 DNA samples belonging to 113 distinct bears selected from the Italian national biobank. We compared data from 4 different marker subsets on the basis of genotyping errors, allelic patterns, observed and expected heterozygosity, discriminatory powers, number of mismatching pairs, and probability of identity. The optimal marker set was selected evaluating the low molecular weight, the high discriminatory power, and the low occurrence of genotyping errors of each primer. We calibrated allele calls and verified matches among genotypes obtained in previous studies using the complete set of 13 STRs (Short Tandem Repeats), analyzing six invasive DNA samples from distinct individuals. Differences in allele-sizing between labs were consistent, showing a substantial overlap of the individual genotyping. CONCLUSIONS: The proposed marker set comprises 11 Ursus specific markers with the addition of cxx20, the canid-locus less prone to genotyping errors, in order to prevent underestimation (maximizing the discriminatory power) and overestimation (minimizing the genotyping errors) of the number of Apennine brown bears. The selected markers allow saving time and costs with the amplification in multiplex of all loci thanks to the same annealing temperature. Our work optimizes the available resources by identifying a shared panel and a uniform methodology capable of improving comparisons between past and future studies.

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.

Does kinship affect spatial organization in a small and isolated population of a solitary felid: The Eurasian lynx?

Social organization in wild carnivores is mostly determined by patterns of family bonds, which may shape the degree of relatedness among individuals in the population. We studied kinship in a small and isolated population of a solitary carnivore, the Eurasian lynx (Lynx lynx) to evaluate its effect on spatial distribution of individuals. We investigated the relationship between spatial location and pair-wise kinship among 28 lynx individuals identified in 2004-2011 by telemetry, non-invasive sampling and genotyping with the use of 12 autosomal microsatellites in the Bialowieza Primeval Forest, Poland. The average relatedness of the lynx population was relatively low (Lynch and Ritland's R = 0.03). Females were significantly more related to each other than males with other males. The inferred pedigree showed that the population was dominated by only 2 familial groups. We did not find significant correlations between the relatedness and the extent of home range overlap or the straight-line distances between the home ranges' central points. These results suggest that the dynamics of kinship in this solitary felid may not differ from the random mating processes described in social carnivores. Although the chances of random mating could be limited to a few resident males and females, the presence of unrelated floaters may provide a "breeding buffer" that may prevent an increase of relatedness and likely inbreeding in the population. This system is likely to fail in preserving genetic diversity in small, highly isolated populations; therefore, restoring habitat connectivity is crucial to ensure sufficient immigration from neighboring populations.