Genes of the extinct Caucasian bison still roam the Białowieża Forest and are the source of genetic discrepances between Polish and Belarusian populations of the European bison,Bison bonasus

European bison (Bison bonasus) populations from both the Polish (PL) and the Belarusian (BY) sides of the Bia?owie?a Forest represent the Lowland genetic line (LB line) – progeny of the Lowland bison (Bison bonasus bonasus) that inhabited western, central, and south‐eastern Europe in historical times. During the species recovery, one of the founders was a descendant of the extinct Caucasian bison (Bison bonasus caucasicus) and its descendants formed the other genetic line – Lowland–Caucasian (LC). There have been justified suspicions that LB European bison in the former Soviet Union had undergone cross‐mating with the LC line. We performed a comparative genetic analyses on European bison from the BY and PL parts of the Bia?owie?a Forest, the LC line and extinct Caucasian bison, based on a set of 19 microsatellite markers and 1512 bovine single nucleotide polymorphism (SNP) markers, polymorphic in at least one of the studied populations. Although genetic variability (mean allele number and expected heterozygosity) for both populations were similar, the F_ST jack‐knifing and principal component analyses PCA revealed highly significant differences between PL and BY bison from the Bia?owie?a Forest. Examining DNA of the extinct Caucasian bison revealed that at least part of the genetic variants found in the BY, but not the PL, population were of Caucasian origin. The results indicate that the contemporary population of European bison from the BY part of the Bia?owie?a Forest should not be regarded as a LB line. The results also suggest that the actual global population size of the LB line European bison is only a half of its official status. Consideration of the presented results are crucial in determining management actions and policy decisions in order to conserve LB line bison within the Bia?owie?a Forest – its natural refuge. © 2015 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, 114 , 752–763.     

Polymorphism of bovine microsatellite DNA sequences in the lowland European bison

Investigations of genetic polymorphism of microsatellite DNA sequences were conducted in 22 individuals of the European bisonBison bonasus (Linneaus, 1758) from Bia?owie?a Primeval Forest. For this purpose 27 cattle microsatellite primer pairs were used. Among the 27 microsatellite markers examined, an amplification product was obtained for 21 loci. This rendered it possible to identify total of 40 alleles in the bison population tested. In addition, eight loci were proved to be monomorphic. A majority of the 40 alleles identified was identical with the alleles identified at the corresponding loci in cattle. Only two alleles seem to be specific for the European bison. The value of heterozygosity for the examined loci in bison population from Bia?owie?a was low and ranged from 0.13 to 0.53. Hence, the polymorphism information content was low as well. Based on our results the microsatellite DNA markers identified in cattle may be used to analyse the genetic structure of the population of European bison.     

Genetic Characteristics and Prognosis for Existence of the European Bison Free-Living Population Created in the Orlovskoe Poles'e National Park

We present preliminary data on genetic aspects of preserving viability of the free-living population of European bison (Bison bonasus L.) created in the Orlovskoe Poles'e National Park.     

Defecation rate in captive European bison, Bison bonasus

Data on the digestive characteristics of European bison, Bison bonasus (L.), are needed for studies of their role as the largest extant herbivore in Europe and a potential keystone species of the temperate forest ecosystem. Very little published data are available, particularly on the defecation rate which affects population estimates from dropping counts and also the individual seed deposition rate. We gathered data from a captive bison group kept at the Show Reserve of the Bia?owie?a National Park. Droppings accumulated in the enclosure over a 72-h period were counted in winter 2010. In addition, the group was observed over approximately 6-h periods three times in winter and 16 times in summer. The count of accumulated droppings over a 72-h period gave eight defecations per day. The summer direct observations recorded 7.5 defecations per day and winter observation 5.4 defecations per day. These estimates are within the range for other bovids of similar size. The difference between summer and winter observation-based estimates may be accounted for by a higher frequency of defecation in early morning and late afternoon, periods not covered in winter observations. Given the published density of seedlings emerging from droppings of the ～470 free-living bison in the nearby forest, eight defecations a day mean that seed deposition by European bison may contribute significantly to realize seed dispersal and plant establishment.     

MHC-DRB3 variation in a free-living population of the European bison, Bison bonasus

MHC genes play a crucial role in pathogen recognition and are the most polymorphic genes in vertebrates. Loss of variation in these genes in bottlenecked species is thought to put their survival at risk. We examined variation at the MHC II DRB3 locus in the European bison, Bison bonasus, a species that has undergone an extreme bottleneck: the current population originated from only 12 founders. We also tested for the association of DRB3 genes with the incidence of posthitis, a disease affecting the reproductive organs of bulls and posing a new threat to the survival of the species. We found very limited MHC diversity, with only four alleles segregating in a sample of 172 individuals from a free‐ranging Białowieża population. The alleles were highly divergent and revealed the hallmark of positive selection acting on them in the past, that is, a significant excess of nonsynonymous substitutions. This excess was concentrated in putative antigen‐binding sites, suggesting that selection was driven by pathogens. However, we did not observe departures from Hardy–Weinberg equilibrium, an indicator of strong ongoing selection. Neither have we found a significant association between DRB3 alleles or genotypes and susceptibility to posthitis. Alleles conferring resistance to males may have been lost during the extreme bottleneck the species had undergone.     

Resting site selection by large herbivores – The case of European bison (Bison bonasus) in Białowieża Primeval Forest

Resting site selection by European bison (Bison bonasus, L., 1758), the largest terrestrial mammal of Europe, was studied in the free-ranging population in Bia?owie?a Primeval Forest (Poland) in 2009–2010. In total, 104 sites of 21 bison (both collared and uncollared) were analysed to determine the most important microhabitat characteristics selected by resting bison during summer and winter and to study the influence of supplementary feeding on resting behaviour of this herbivore. Resting sites were identified on the basis of GPS locations and activity records collected by GPS collars, as well as direct observations of bison, and were compared with control sites. Microhabitat selection by bison did not differ significantly between the sexes. During summer and winter, bison resting sites displayed a high tree density, low visibility and high complexity (structures providing cover). Summer resting sites were also characterised by a significantly lower abundance of blood-sucking insects and denser canopy than control sites. Winter resting sites showed a lower complexity and higher visibility than summer sites, and were less often located in mixed forest habitats. During winter, bison rested more frequently in forest below 50 years of age than in older forest. Resting sites of non-fed bison were more often located in young coniferous forests, were lower in visibility and situated closer to open areas than sites of bison using supplementary winter feeding, suggesting a trade-off between food and cover. The results indicate that European bison select their resting sites in areas of mosaic habitat structure providing cover from disturbances with access to profitable natural forage grounds.     

Phylogenetic relationships among the European and American bison and seven cattle breeds reconstructed using the BovineSNP50 Illumina Genotyping BeadChip

Here we present the first attempt to use the BovineSNP50 Illumina Genotyping BeadChip for genome-wide screening of European bison Bison bonasus bonasus (EB), two subspecies of American bison: the plains bison Bison bison bison (PB), the wood bison Bison bison athabascae (WB) and seven cattle Bos taurus breeds. Our aims were to (1) reconstruct their evolutionary relationships, (2) detect any genetic signature of past bottlenecks and to quantify the consequences of bottlenecks on the genetic distances amongst bison subspecies and cattle, and (3) detect loci under positive or stabilizing selection. A Bayesian clustering procedure (STRUCTURE) detected ten genetically distinct clusters, with separation among all seven cattle breeds and European and American bison, but no separation between plain and wood bison. A linkage disequilibrium based program (LDNE) was used to estimate the effective population size (N _e) for the cattle breeds; N _e was generally low, relative to the census size of the breeds (cattle breeds: mean N _e = 299.5, min N _e = 18.1, max N _e = 755.0). BOTTLENECK 1.2 detected signs of population bottlenecks in EB, PB and WB populations (sign test and standardized sign test: p = 0.0001). Evidence for loci under selection was found in cattle but not in bison. All extant wild populations of bison have shown to have survived severe bottlenecks, which has likely had large effects on genetic diversity within and differentiation among groups.     

Effects of a recent founding event and intrinsic population dynamics on genetic diversity in an ungulate population

Maintenance of genetic diversity has recently become a management goal for a number of species, due to its importance for present and future population viability. Genetic drift, primarily through differential reproductive success and inbreeding, can accelerate the loss of genetic diversity in recently recovered populations. We attempt to quantify the consequences of these factors on the genetic diversity contained in a small, recently founded wood bison (Bison bison athabascae) population by examining the genetic variation in this conservation herd, the calves born therein, and its large source population. The Hook Lake Wood Bison Recovery Project was initiated to found a disease-free herd of wood bison containing a representative amount of the genetic diversity present in the Wood Buffalo National Park metapopulation. Levels of diversity in the Hook Lake Wood Bison Recovery Project founders are higher than in previous salvage attempts. To examine the effects of differential reproductive success on this population, we monitored parentage of the calves born in the Hook Lake Wood Bison Recovery Project for 3 years since the founders reached sexual maturity. Two of the male founders sired over 90% of the offspring born in this population, which has led to a reduction in diversity in their calves. Monitoring of reproductive success, and incorporation of selective breeding strategies will be required to reduce the rate at which genetic diversity is lost from this small, isolated population. These steps should occur in other recovery projects, particularly when a small number of individuals are capable of dominating reproduction.     

Genetic variability in the European bison

In the European bison, Bison bonasus (L), 15 proteins coded by 20 loci have been studied. Two of these loci (Es-3 and Ca) are polymorphic and the remaining 18 are monomorphic. The average heterozygosity of bisons in Polish herds is 3.5%. The value does not differ from those found in populations of other large mammals in spite of restoration of the European bison population from a mere 13 individuals.     

Movements of European bison (Bison bonasus) beyond the Białowieża Forest (NE Poland): range expansion or partial migrations?

The European bison is the largest terrestrial mammal in Europe. After extinction in the wild at the beginning of the twentieth century, it was re-introduced to Bia?owie?a Forest and other woodlands in Eastern Europe. In this paper, we analysed the movements of European bison beyond the continuous woodlands of the Bia?owie?a Forest (NE Poland) between 1964 and 2010. In total, 1,117 direct observations of bison were collected. The number of males moving out of the forest fluctuated during the study period, whilst the number of females steadily increased. The number of male observations outside of the forest per annum was dependent on the population size and snow depth, whilst the number of cows in mixed groups moving outside of the forest was correlated with the population size only. Males were observed mainly alone (50 % of observations) or in small groups of two to three individuals (25 %); however, distribution of group size differed from those observed in the population. There was a significant difference between the direction of movement of males and females out of the forest—males moved mainly west and southwest, whilst females moved to the north. This was also significantly different from the expected movement direction. The mean distance of bison observations from the forest border was 1.8?±?0.13 km and did not differ significantly between sexes. After 1990, males were observed significantly farther away from the forest (2.2 km) than in the previous years (0.9 km). Most observations (94 % of bulls and 93 % of cows) were up to 5 km from the forest edge. The range of bison in the vicinity of the Bia?owie?a Forest was strictly seasonal. Most observations (78 % in males and 88 % in females) were recorded from November to April. Increasing utilisation of areas beyond the forest habitats may be driven by different factors but most probably it is related to range expansion and the bison's preference for open habitats. The strong seasonal pattern of bison movements indicate that the partial seasonal migrations were initiated in the Bia?owie?a population.     

Antibodies against Mycoplasma bovigenitalium in free-living European bison (Bison bonasus) with balanoposthitis

Since 1980 severe chronic balanoposthitis has been observed in free-living European bison (Bison bonasus) in the Bia?owieza Primeval Forest (Poland). Sera of 50 bison with balanoposthitis and 48 clinically healthy male and 49 female bison were investigated for antibodies against Mycoplasma bovis and M. bovigenitalium by western blot analysis. Prevalence of antibodies against M. bovigenitalium was significantly higher in bison with balanoposthitis than in unaffected male bison. Mycoplasma bovigenitalium may play a role in the pathogenesis of balanoposthitis in European bison.     

Did the historical range of the European bison (<Emphasis Type="Italic">Bison bonasus</Emphasis> L.) extend further south?—a new finding from the Yenikapı Metro and Marmaray excavation,Turkey

The origin of the European bison (Bison bonasus, Linnaeus, 1758) has been widely discussed and investigated in recent years. The species had a wide historic geographic distribution throughout the European continent during the middle and late Holocene, ranging from France in the west to the Caucasus in the east. However, archaeological evidence is needed to resolve the southern extent of the European bison distribution. We discovered one bison skull fragment during archaeological excavations in 2008 in the area of Yenikap? Metro and Marmaray (Turkey). Radiocarbon dating indicated the skull was deposited during the Byzantine period (seventh to eighth century AD). Mitochondrial genome analyses provided clear evidence that the skull was from a European bison. This is the first unambiguous evidence of the presence of this species in southeastern Europe during Byzantine times, which validates the historical written records of a potentially wider range of the European bison in historical times.     

Genetic variation within and relatedness among wood and plains bison populations.

There are two recognized subspecies of bison, wood (Bison bison athabascae) and plains (Bison bison bison) bison. The establishment of most bison populations from a small number of individuals has raised concerns about their genetic variation. To this end, 11 bison populations were surveyed with 11 microsatellite loci in order to calculate genetic variation and genetic distances. Mean number of alleles ranged between 3.18 at Antelope Island State Park (Utah) and 6.55 at Wood Buffalo National Park (Alberta and Northwest Territories). Mean heterozygosity ranged from 0.295 at Antelope Island State Park to 0.669 at Custer State Park (South Dakota). The amount of genetic variability present in the bison populations as measured by mean number of alleles and overall probability of identity was found to correlate with the number of founders for all sampled populations. The G-test for heterogeneity revealed some evidence for the existence of subpopulations at Wood Buffalo National Park, however very small genetic distances between these subpopulations suggest that nuclear material from the plains bison introduced into Wood Buffalo National Park has diffused throughout the park. Genetic distances between the sampled populations were generally larger between than within the two bison subspecies.     

Deletion/insertion polymorphism of the prion protein gene (<Emphasis Type="Italic">PRNP</Emphasis>) in Polish Red cattle,Polish White-backed cattle and European bison (<Emphasis Type="Italic">Bison bonasus</Emphasis> L., 1758)

The aim of the present study was to identify deletion/insertion polymorphism of the bovine prion protein (PRNP) gene within the promoter sequence (23 bp indel), intron 1 (12 bp indel) and the 3′ end untranslated region (14 bp indel). The experiment was performed on three groups of animals protected under a genetic resources conservation program: 139 Polish Red (PR) cows, 79 Polish White-backed cows and 50 European bison (Bison bonasus L., 1758). White-backed cattle were characterized by a higher frequency of ins/del heterozygotes and a relatively lower frequency of ins/ins homozygotes within the promoter sequence region (23 bp indel), compared to Polish Red cattle. At the polymorphic locus of intron 1 (12 bp indel) the genetic structure of both cattle populations was similar. Monomorphism, expressed by the occurrence of one genotype variant in each of the analyzed sequence regions, was observed in European bison. Five haplotypes were found in Polish White-backed cows, four haplotypes in Polish Red cows and only one in analyzed group of bison. Differences between the observed and expected number of PRNP haplotypes were recorded in Polish Red cattle. The article is published in the original.     

Body mass and measurements of the European bison during postnatal development

Body mass of the European bisonBison bonasus (Linnaeus, 1758) was determined for 152 animals (86 males, 66 females) from captive breeding centre in Białowieża and for 492 (248, 244) animals from the free-ranging herd in the Białowieża Forest. Body measurements were taken for a total of 472 bison (204, 268). Age-related changes in body mass and measurements of 375 bison (165, 210) were analysed. Bison body mass was significantly correlated with age. Captive bison were heavier than those from a free-ranging herd. Measurements showed the changes in bison body structure in the postnatal development. All measurements were highly correlated with age and mass of the animals and increased most intensively during the first year of life. Sex-related differences in body mass and measurements were most pronounced in bison ≥ 3 years old. Physical development of females ends at the age of 5 years, and that of males at the age of 7 years.     

New data on the evolutionary history of the European bison (Bison bonasus) based on subfossil remains from Southeastern Europe

The origin and evolutionary history of the European bison Bison bonasus (wisent) have become clearer after several morphological, genomic, and paleogenomic studies in the last few years, but these paleogenomic studies have raised new questions about the evolution of the species. Here, we present additional information about the population diversity of the species based on the analysis of new subfossil Holocene remains from the Balkan Peninsula. Seven ancient samples excavated from caves in Western Stara Planina in Bulgaria were investigated by mitochondrial D‐loop (HVR1) sequence analysis. The samples were dated to 3,800 years BP by radiocarbon analysis. Additionally, a phylogenetic analysis was performed to investigate the genetic relationship among the investigated samples and all mitochondrial DNA sequences from the genus Bison available in GenBank. The results clustered with the sequences from the extinct Holocene South‐Eastern (Balkan) wisent to the fossil Alpine population from France, Austria, and Switzerland, but not with those from the recent Central European (North Sea) one and the now extinct Caucasian population.In conclusion, these data indicate that the Balkan wisent that existed in historical time represented a relict and probably an isolated population of the Late Pleistocene‐Holocene South‐Western mountainous population of the wisent.     

Fluctuating ungulate density shapes tree recruitment in natural stands of the Białowieża Primeval Forest,Poland

Question: What are the main driving factors in 70 years of natural dynamics in tree recruitment in the Bia?owie?a National Park? Location: Bia?owie?a National Park, Poland, is one of the least disturbed temperate, lowland forest systems in Europe. Methods: We tested whether fluctuations in large herbivore populations, changes in climate and openness of the forest explained compositional dynamics. Tree recruitment (to size class DBH≥5 cm) was measured on permanent transects (in total, 14.9 ha) six times between 1936‐2002. These data were related to existing data on ungulate density, climatic parameters and estimates of forest openness collected during the same period. Results: Total recruitment of all tree species combined was negatively correlated with total ungulate density and red deer density. The variation in response between species was related to the preferences of herbivores; the more preferred forage species (especially Carpinus betulus) were positively and the less preferred species negatively related to herbivore density. Total tree recruitment rates were not related to climatic parameters and openness of the forest. Only Alnus glutinosa recruitment was significantly related to climatic parameters, and Ulmus glabra related to forest openness, but there were no predictable patterns in recruitment among species in relation to these factors. Conclusion: The present study indicated that changes in large herbivore density have played an important role in driving patterns in tree recruitment and species composition during the last 70 years in Bia?owie?a National Park. In contrast to other studies, increasing herbivore numbers were associated with higher recruitment of preferred and browsing‐tolerant species. Periodical crashes in ungulate numbers, whether human‐induced or caused by natural factors, may offer windows of opportunity for regeneration of a range of tree species and facilitate more diverse and dynamic forest development.     

Adapt or die—Response of large herbivores to environmental changes in Europe during the Holocene

Climate warming and human landscape transformation during the Holocene resulted in environmental changes for wild animals. The last remnants of the European Pleistocene megafauna that survived into the Holocene were particularly vulnerable to changes in habitat. To track the response of habitat use and foraging of large herbivores to natural and anthropogenic changes in environmental conditions during the Holocene, we investigated carbon (δ¹³C) and nitrogen (δ¹⁵N) stable isotope composition in bone collagen of moose (Alces alces), European bison (Bison bonasus) and aurochs (Bos primigenius) in Central and Eastern Europe. We found strong variations in isotope compositions in the studied species throughout the Holocene and diverse responses to changing environmental conditions. All three species showed significant changes in their δ¹³C values reflecting a shift of foraging habitats from more open in the Early and pre‐Neolithic Holocene to more forest during the Neolithic and Late Holocene. This shift was strongest in European bison, suggesting higher plasticity, more limited in moose, and the least in aurochs. Significant increases of δ¹⁵N values in European bison and moose are evidence of a diet change towards more grazing, but may also reflect increased nitrogen in soils following deglaciation and global temperature increases. Among the factors explaining the observed isotope variations were time (age of samples), longitude and elevation in European bison, and time, longitude and forest cover in aurochs. None of the analysed factors explained isotope variations in moose. Our results demonstrate the strong influence of natural (forest expansion) and anthropogenic (deforestation and human pressure) changes on the foraging ecology of large herbivores, with forests playing a major role as a refugial habitat since the Neolithic, particularly for European bison and aurochs. We propose that high flexibility in foraging strategy was the key for survival of large herbivores in the changing environmental conditions of the Holocene.     

Morphometric variation of the skull during postnatal development in the Lowland European bisonBison bonasus bonasus

We studied the variation of linear measurements and skull capacity in Lowland European bisonBison bonasus bonasus (Linnaeus, 1758) during postnatal development, and the dependencies of the parameters in relation to sex, age, and body mass of the animals. Material consisted of 599 bison skulls (310 males and 289 females), within the age range of 1 month to 21 years (males) and to 27 years (females). In the group of calves to 1 year old, no sex connected differences in skull measurements were observed, whereas the skull capacity in older calves was significantly larger (0.01>p>0.001) in males than in females. From the third year of life, most skull measurements display characteristics of sexual dimorphism. Skull development in both sexes is most intensive during the first three years of life, and slows from the age of 5. In older individuals of both sexes (≥ 6 years), orbital breadth continues growing and, in females, breadth of splanchnocranium continues increasing. Growth in a bison’s skull capacity is most intensive up to the third year of life and slows from the age of 5. During postnatal development, a bison skull grows proportionally except the neurocranium, which grows slightly slower in comparison with basal length and its development finishes earlier than that of splanchnocranium. In ontogenesis, a bison skull grows much slower compared to body mass. In relation to body mass, skull capacity and the height of neurocranium grow most slowly while orbital breadth grows most intensively. The results obtained were compared with data on skull sizes of bison born in 1930–1950 and bred in captivity and with skulls of the American bisonBison bison. Inbreeding is probably responsible for some types of phenotypic abnormalities in the skull which appear in modern European bison.