Alexandros A. Karamanlidis

North-south differentiation and a region of high diversity in European wolves (Canis lupus)

European wolves (Canis lupus) show population genetic structure in the absence of geographic barriers, and across relatively short distances for this highly mobile species. Additional information on the location of and divergence between population clusters is required, particularly because wolves are currently recolonizing parts of Europe. We evaluated genetic structure in 177 wolves from 11 countries using over 67K single nucleotide polymorphism (SNP) loci. The results supported previous findings of an isolated Italian population with lower genetic diversity than that observed across other areas of Europe. Wolves from the remaining countries were primarily structured in a north-south axis, with Croatia, Bulgaria, and Greece (Dinaric-Balkan) differentiated from northcentral wolves that included individuals from Finland, Latvia, Belarus, Poland and Russia. Carpathian Mountain wolves in central Europe had genotypes intermediate between those identified in northcentral Europe and the Dinaric-Balkan cluster. Overall, individual genotypes from northcentral Europe suggested high levels of admixture. We observed high diversity within Belarus, with wolves from western and northern Belarus representing the two most differentiated groups within northcentral Europe. Our results support the presence of at least three major clusters (Italy, Carpathians, Dinaric-Balkan) in southern and central Europe. Individuals from Croatia also appeared differentiated from wolves in Greece and Bulgaria. Expansion from glacial refugia, adaptation to local environments, and human-related factors such as landscape fragmentation and frequent killing of wolves in some areas may have contributed to the observed patterns. Our findings can help inform conservation management of these apex predators and the ecosystems of which they are part.

Noninvasive genetic studies of brown bears using power poles

One difficulty in the conservation of endangered wildlife is the lack of reliable information on its status. This lack of knowledge can often be attributed to financial and logistic constraints as well as the lack of trained personnel to collect data. We test a simple method to study bears in the southern Balkans by inspecting power poles, which are used by bears for marking and rubbing purposes. We created a network of barbed-wire fitted poles for the collection of hair samples, evenly distributed throughout six study areas. During 87 sampling sessions in the main study area, we collected 191 samples and identified six microsatellite loci that were variable enough for individual bear identification. The most and best-quality hair samples were collected during the mating period, and DNA was most successfully extracted from samples remaining <4xa0weeks in the field. In the six study areas, we identified 47 bears. An advantage of using power poles for hair sampling is their availability and accessibility; no bait is required, and the network can be easily set up. A drawback may be an unequal capture probability of sex and age classes of bears. Despite this limitation, using power poles proved to be a simple and cheap method for the noninvasive genetic study of bears that did not require any prior knowledge on habitat use and activity patterns. The method is suitable for large-scale surveys to estimate distribution and relative densities of bears and could also be applied for studying other species.

Using sign at power poles to document presence of bears in Greece

Abstract The endangered brown bear (Ursus arctos) population in Greece is in urgent need of effective protection and management; that management should be based on information that is both reliable and quickly attained. After observing bears marking and rubbing on power poles, we initiated a study to collect information on this behavior and develop an effective method for documenting bear presence in Greece. Thirty-nine power poles in the main study area were fitted with barbed wire and inspected monthly for a year. The information and experience gained in the main study area was used to survey 3 additional areas, covering a representative sample of the species distribution in the country. Power pole-related behaviors were associated with mud smears, hair deposits, and bite and claw marks (hereafter referred to as marks). Tracks and scats also have been used to document the presence of brown bears in Greece, but fewer of these were found in all areas surveyed. Deterioration rate of marks was slower than that of tracks and scats. Our results suggest that power pole-related behavior is not a localized phenomenon. A monitoring scheme in Greece documenting the presence of the species that would include the regular inspection of power poles could take advantage of the higher abundance and slower deterioration rate of power pole-related signs and be time efficient and easily staffed by volunteers. The ability to identify individual bears through genetic analysis of hair collected from power poles is an additional advantage of this approach.

The availability of resting and pupping habitat for the Critically Endangered Mediterranean monk seal Monachus monachus in the archipelago of Madeira

In order to describe the resting and pupping habitat of the Critically Endangered Mediterranean monk seal Monachus monachus and facilitate the identification of preferences by the species for suitable habitat for resting and pupping, 94 sea caves in the archipelago of Madeira were located, charted, and categorized using six characteristics. A cluster analysis indicated that there are eight different types of cave, in three groups. Observations of cave usage indicate that monk seals in the archipelago do not appear to exhibit a preference for certain cave types for resting. When taking care of young, however, the species appears to prefer sea caves that have beaches above sea level during high tide and long entrance corridors. Based on these preferences we believe that although there is a large number of caves in the area that are suitable for resting, only 16 (17%) of the caves are suitable for pupping. The survival of the Critically Endangered Mediterranean monk seal will depend on the allocation of sufficient suitable habitat for reproduction.

Genetic diversity, structure, and size of an endangered brown bear population threatened by highway construction in the Pindos Mountains, Greece

One of the major negative effects of roads is the creation of barriers to the movement of wildlife, ultimately disconnecting populations and increasing extinction risk. We collected genetic data from a threatened brown bear population in the central part of the Pindos mountain range in northwestern Greece to provide information about this, as yet genetically undescribed, population and to evaluate its status prior to the construction of a major highway. We used noninvasive genetic sampling methods and microsatellite analysis to investigate nuclear genetic diversity, population genetic structure, demographic history, relatedness within the population and estimated effective and total population size. Brown bears in the study area were found to possess a relatively high level of nuclear genetic diversity and low levels of inbreeding; the population did not show any signs of substructuring but seems to have gone through a genetic bottleneck in the recent past. The estimated effective population size was 29, and the total population size estimate obtained by two different methods was 33 and 51 individuals, respectively. Our results indicate a good conservation status of this bear population and provide baseline genetic data for the future evaluation of the effects on bears from the construction of a major highway, for monitoring the genetic status of this and other bear populations in Greece and for assessing gene flow in bear populations in southern Europe.

Structural and economic aspects of human–bear conflicts in Greece

Abstract Agricultural damage and the resulting negative attitudes of farmers are major issues in the conservation of brown bears (Ursus arctos). We analyzed 3,241 approved compensation claims to gain insight into human–brown bear conflicts in Greece from 1999 to 2006. Damage to livestock was low compared to the number of livestock in an area and affected mainly young cattle and single equids. Damage to sheep was low in Greece in comparison to other countries. Crop damage was recorded mainly in small corn fields and vineyards, while damage to apiaries was associated with their general availability in an area and resulted in considerable economic losses. Bear damage occurred throughout the year, but was most common from May to October, and with the exception of crop damage, was correlated with the current range of the species. To decrease damage levels by bears in Greece and considering the current management and conservation circumstances for the species in the country, we propose the large-scale promotion and use of livestock guarding dogs and electric fencing for small fields of valuable crops as well as apiaries. Reduction of depredation to cattle will require structural changes to the way herds are managed, and compensation for damage should be linked to active damage prevention. On a local scale the livestock husbandry systems may be adjusted by increasing herd size and by penning vulnerable livestock overnight.

Monitoring Human Activity in an Area Dedicated to the Protection of the Endangered Mediterranean Monk Seal

Mediterranean monk seal populations have declined dramatically and survive in subpopulations scattered throughout their original distribution. The National Marine Park of Alonnisos, Northern Sporades is the first marine protected Area dedicated to the protection of the species in the Mediterranean. A Monitoring and Awareness Project has been carried out in the area from 1993 to 2000, the results of which have produced a detailed profile of the human usage pattern of the Park and indicated a significant reduction of illegal activities in the area. The pattern of human usage has enabled the monitoring team to adjust their efforts in the Park over time and provided essential baseline data for future management. The combined monitoring and awareness efforts have contributed significantly to the conservation of the Mediterranean monk seal by successfully protecting pupping sites and reducing human-induced mortality of the species in the area.

Genome‐wide analyses suggest parallel selection for universal traits may eclipse local environmental selection in a highly mobile carnivore

Abstract Ecological and environmental heterogeneity can produce genetic differentiation in highly mobile species. Accordingly, local adaptation may be expected across comparatively short distances in the presence of marked environmental gradients. Within the European continent, wolves (Canis lupus) exhibit distinct north–south population differentiation. We investigated more than 67‐K single nucleotide polymorphism (SNP) loci for signatures of local adaptation in 59 unrelated wolves from four previously identified population clusters (northcentral Europe n = 32, Carpathian Mountains n = 7, Dinaric‐Balkan n = 9, Ukrainian Steppe n = 11). Our analyses combined identification of outlier loci with findings from genome‐wide association study of individual genomic profiles and 12 environmental variables. We identified 353 candidate SNP loci. We examined the SNP position and neighboring megabase (1 Mb, one million bases) regions in the dog (C. lupus familiaris) genome for genes potentially under selection, including homologue genes in other vertebrates. These regions included functional genes for, for example, temperature regulation that may indicate local adaptation and genes controlling for functions universally important for wolves, including olfaction, hearing, vision, and cognitive functions. We also observed strong outliers not associated with any of the investigated variables, which could suggest selective pressures associated with other unmeasured environmental variables and/or demographic factors. These patterns are further supported by the examination of spatial distributions of the SNPs associated with universally important traits, which typically show marked differences in allele frequencies among population clusters. Accordingly, parallel selection for features important to all wolves may eclipse local environmental selection and implies long‐term separation among population clusters.

Distribution and genetic status of brown bears in FYR Macedonia: implications for conservation

Conservation and management of large carnivores is often hampered by the lack of information of basic biological parameters. This is particularly true for brown bears (Ursus arctos) in the Former Yugoslav Republic (FYR) of Macedonia. The bear population in this country is important, as it links bear populations of the central part of the Dinaric–Pindos population and the endangered population to the south in Greece. The aim of this study was to assess bear presence in FYR Macedonia and to provide the first evaluation of the genetic status of the species in this country. Bear presence was assessed through a questionnaire and sign surveys, while the genetic status of the species was evaluated through noninvasive genetic sampling from power poles and microsatellite analysis. The results of the study indicate the continuous and permanent presence of brown bears in FYR Macedonia from the border to Kosovo in the northwest, along the border to Albania and Greece in the south; bear presence around Mount Kožuf in the south of the country was seasonal. High levels of genetic diversity were recorded, and it appears that this bear population is currently not threatened by low genetic variability. Cross-border movements of bears between FYR Macedonia and Greece were documented, indicating the presence of an interconnected population and outlining the necessity for a coordinated international approach in the monitoring and conservation of the species in southeastern Europe.

Survival and divergence in a small group: The extraordinary genomic history of the endangered Apennine brown bear stragglers

Significance A small and relict population of brown bears lives in complete isolation in the Italian Apennine Mountains, providing a unique opportunity to study the impact of drift and selection on the genomes of a large endangered mammal and reconstruct the phenotypic consequences and the conservation implications of such evolutionary processes. The Apennine bear is highly inbred and harbors very low genomic variation. Several deleterious mutations have been accumulated by drift. We found evidence that this is a consequence of habitat fragmentation in the Neolithic, when human expansion and land clearance shrank its habitat, and that retention of variation at immune system and olfactory receptor genes as well as changes in diet and behavior prevented the extinction of the Apennine bear. About 100 km east of Rome, in the central Apennine Mountains, a critically endangered population of ∼50 brown bears live in complete isolation. Mating outside this population is prevented by several 100 km of bear-free territories. We exploited this natural experiment to better understand the gene and genomic consequences of surviving at extremely small population size. We found that brown bear populations in Europe lost connectivity since Neolithic times, when farming communities expanded and forest burning was used for land clearance. In central Italy, this resulted in a 40-fold population decline. The overall genomic impact of this decline included the complete loss of variation in the mitochondrial genome and along long stretches of the nuclear genome. Several private and deleterious amino acid changes were fixed by random drift; predicted effects include energy deficit, muscle weakness, anomalies in cranial and skeletal development, and reduced aggressiveness. Despite this extreme loss of diversity, Apennine bear genomes show nonrandom peaks of high variation, possibly maintained by balancing selection, at genomic regions significantly enriched for genes associated with immune and olfactory systems. Challenging the paradigm of increased extinction risk in small populations, we suggest that random fixation of deleterious alleles (i) can be an important driver of divergence in isolation, (ii) can be tolerated when balancing selection prevents random loss of variation at important genes, and (iii) is followed by or results directly in favorable behavioral changes.