Detection of a newly described pestivirus of Pyrenean chamois (Rupicapra pyrenaica pyrenaica) in France

A pestivirus was detected and characterized in chamois (Rupicapra pyrenaica pyrenaica) originating from the French part of the Pyrenees. Phylogenetic analysis of the pestivirus was done on the basis of a fragment from the 5' noncoding region including 22 published nucleotide sequences of different pestivirus strains. Our strain was grouped within the clade of border disease viruses (BDV). However, it had an intermediate position between clade BDV and classical swine fever viruses representing a basal position to BDV strains of domestic sheep. Our strain was grouped as a sister unit to a novel pestivirus (Chamois-1) recently described from chamois in Spain. Therefore, we postulate that this virus occurs in the entire population of Pyrenean chamois. On the basis of the phylogenetic grouping of this isolate, a postulated cross-species transmission of pestivirus from domestic sheep to chamois via shared pastures seems to be unlikely.     

Diversity and evolution of the Mhc-DRB1 gene in the two endemic Iberian subspecies of Pyrenean chamois, Rupicapra pyrenaica

Major histocompatibility complex class II locus DRB variation was investigated by single-strand conformation polymorphism analysis and sequence analysis in the two subspecies of Pyrenean chamois (Rupicapra pyrenaica) endemic to the Iberian Peninsula. Low levels of genetic variation were detected in both subspecies, with seven different alleles in R. p. pyrenaica and only three in the R. p. parva. After applying the rarefaction method to cope with the differences in sample size, the low allele number of parva was highlighted. The low allelic repertoire of the R. p. parva subspecies is most likely the result of bottlenecks caused by hunting pressure and recent parasitic infections by sarcoptic mange. A phylogenetic analysis of both Pyrenean chamois and DRB alleles from 10 different caprinid species revealed that the chamois alleles form two monophyletic groups. In comparison with other Caprinae DRB sequences, the Rupicapra alleles displayed a species-specific clustering that reflects a large temporal divergence of the chamois from other caprinids, as well as a possible difference in the selective environment for these species.     

Vigilance behavior of Pyrenean chamois Rupicapra pyrenaica pyrenaica: Effect of sex and position in the herd

The Pyrenean chamois Rupicapra pyrenaica pyrenaica is a mountain-dwelling ungulate with an extensive presence in open areas. Optimal group size results from the trade off between advantages (a reduction in the risk of predation) and disadvantages (competition between members of the herd) of group living. In addition, advantages and disadvantages of group living may vary depending on the position of each individual within the herd. Our objective was to study the effect of central vs. Peripheral position in the herd on feeding and vigilance behavior in male and female Pyrenean chamois and to ascertain if a group size effect existed. We used focal animal sampling and recorded social interactions when a focal animal was involved. With males, vigilance rate was higher in the central part of the group than at the periphery, probably due to a higher density of animals in the central part of the herd and a higher probability of being disturbed by conspecifics. With females, vigilance rate did not differ according to position in the herd. Females spent more time feeding than males, and males showed a higher frequency of the vigilance behavior than females. We did not observe a clear relationship between group size and vigilance behavior. The differences in vigilance behavior might be due to social interactions.     

Predicting seasonal and spatial variations in diet quality of Pyrenean chamois (Rupicapra pyrenaica pyrenaica) using near infrared reflectance spectroscopy

Monitoring nutritional status of wildlife is key to understanding the population response to changes in food availability. Several direct and indirect methods have been proposed for such purposes, but faecal nitrogen (FN) is by far the most commonly used indirect non-invasive indicator of free-ranging ruminant diet quality. In this work, two methods were used to estimate the FN content in 291 pellet samples of Pyrenean chamois (Rupicapra pyrenaica pyrenaica) collected during a 2-year period (May 2009 to May 2011) in two chamois populations from the Catalan Pyrenees, Spain: the Dumas LECO analyzer and near-infrared reflectance spectroscopy (NIRS). In order to optimize further FN estimations we performed three different NIRS calibrations: (1) general, across year, season and populations; (2) population-specific; and (3) season-specific, i.e., different periods of vegetation growth. FN ranged from 1.66 % to 2.85 % (dry matter basis) and was higher in the main vegetation growing season than in winter. Concerning NIRS calibration, our study confirmed that FN of Pyrenean chamois can be accurately determined by NIRS, since the general calibration was accurate enough to predict FN. It is concluded that NIRS represents a non-destructive, emission-free and low-cost analytical technique which may reduce the use of conventional laborious methods for estimating FN in long-term wildlife monitoring.     

Two Different Epidemiological Scenarios of Border Disease in the Populations of Pyrenean chamois (Rupicapra p. pyrenaica) after the First Disease Outbreaks

Since 2001 several outbreaks of a new disease associated with Border disease virus (BDV) infection have caused important declines in Pyrenean chamois (Rupicapra pyrenaica) populations in the Pyrenees. The goal of this study was to analyze the post-outbreak BDV epidemiology in the first two areas affected by disease with the aim to establish if the infection has become endemic. We also investigated if BDV infected wild and domestic ruminants sharing habitat with chamois. Unexpectedly, we found different epidemiological scenarios in each population. Since the disease outbreaks, some chamois populations recuperated quickly, while others did not recover as expected. In chamois from the first areas, prevalence was high (73.47%) and constant throughout the whole study period and did not differ between chamois born before and after the BDV outbreak; in all, BDV was detected by RT-PCR in six chamois. In the other areas, prevalence was lower (52.79%) and decreased during the study period; as well, prevalence was significantly lower in chamois born after the disease outbreak. No BDV were detected in this population. A comparative virus neutralisation test performed with four BDV strains and one Bovine viral diarrhoea virus (BVDV) strain showed that all the chamois had BDV-specific antibodies. Pestivirus antibodies were detected in all the rest of analyzed species, with low prevalence values in wild ruminants and moderate values in domestic ruminants. No viruses were detected in these species. These results confirm the hypothesis that outbreaks of BDV infection only affect the Pyrenean chamois, although other wild ruminants can occasionally be infected. In conclusion, two different scenarios have appeared since the first border disease outbreaks in Pyrenean chamois: on the one hand frequent BDV circulation with possible negative impact on population dynamics in some areas and on the other, lack of virus circulation and quick recovery of the chamois population.     

Social Interactions in Isard Kids Rupicapra pyrenaica pyrenaica (Artiodactyla: Bovidae): Possible Influence on Development of Social Bonding

Assuming that social experiences at an early age may be a determinant factor in the process of formation of social network and attachments, we studied the social interactions of isard, or Pyrenean chamois, kids (Rupicapra pyrenaica pyrenaica) during their first year, to understand “how” sociality arises in isards rather than “why”. Interactions with the mother were affiliative, and approaches of kid to mother or vice versa were generally followed by another “positive” interaction. Interactions between kids were also frequent and playful. Interactions with both mother and peers provided positive reinforcement which may strengthen bonds. However, early interactions of kids with yearlings and other adults provided negative experience, no attachment thus developing from them. Social affinities of adults as well as the social and spatial organization of groups may be understood when ontogenetical history and immediate causality are highlighted.     

Seasonal Changes in Fecal Cortisol Metabolites in Pyrenean Chamois

Abstract: We studied seasonal changes in fecal Cortisol metabolites (FCM), which have been widely used as indicators of stress, in a population of Pyrenean chamois (Rupicapra pyrenaica pyrenaica) in the Cadí Range of northeastern Spain. We collected fecal samples from 2001 to 2003 in 3 particular locations with different altitudes and male or female presence, and we analyzed them for FCM and fecal nitrogen as an indicator of diet quality. We observed a clear seasonal pattern, with the highest FCM in winter, and we obtained correlations between FCM and monthly mean minimum temperatures and fecal nitrogen. We observed no effects of tourism presence, trophy hunting, or rut season on FCM. Analysis of cortisol metabolites in feces can be a good measure of winter stress in Pyrenean chamois.     

Mineral Levels in Pyrenean Chamois (Rupicapra pyrenaica)

The effects of mineral deficiencies are often sub-clinical, and the importance of mineral status is often underestimated in wildlife populations. To our knowledge, this is the first study that gives reference intervals of hepatic minerals for Pyrenean chamois (Rupicapra pyrenaica). We determined macro and trace mineral concentrations in liver samples from 100 animals (44 healthy and 56 sick) collected in the Catalan Pyrenees (NE Spain) from 1995 to 2008. After wet digestion, we determined Na, K, Ca, P, Mg, S, and Fe concentrations by inductively coupled plasma optical emission spectrometry (ICP-OES) and Cu, Zn, and Mo concentrations by inductively coupled plasma mass spectrometry (ICP-MS). We observed low hepatic concentrations of Cu in a considerable percentage of chamois, without evidences that these low concentrations increased their susceptibility to infectious diseases. The group of sick chamois had very similar percentage of animals (10/56) with low concentration of Cu (<20 ppm DW) than the group of healthy chamois (9/44). On the other hand, we observed that infectious diseases increased significantly the hepatic concentrations of Na, Ca, Mg, Fe and Zn, very likely, as a consequence of processes associated with the acute phase inflammatory response. The obtained values of liver mineral levels and their sources of variation, such as sex, age and disease, mostly fall within the range of those described for other ruminants, but possible deficiencies and differences between individuals and populations require further study.     

Prevalence of antibodies against selected agents shared between Cantabrian chamois (Rupicapra pyrenaica parva) and domestic goats

Southern chamois (Rupicapra pyrenaica) share the habitat with domestic ungulates, and may, therefore, play a role in the epidemiology of shared agents. The objective of this study was to determine the seroprevalence for Brucella spp., Mycobacterium avium ssp. paratuberculosis (MAP), pestivirus, and Sarcoptes scabiei in Cantabrian chamois (Rupicapra pyrenaica parva) and compare these data with those of sympatric domestic goats (Capra hircus). From 2005 to 2008, blood samples were obtained from 236 adult Cantabrian chamois in two different populations, the western one and the eastern one. Seroprevalence for Brucella spp. and pestivirus was assessed using commercial ELISA kits, whereas specifically designed ELISA tests were used for MAP and S. scabiei. No antibodies against Brucella spp. were detected. Conversely, antibodies against MAP, pestivirus (chamois 3.8%; goat 2.3%), and S. scabiei (chamois 11.9%; goat 12.8%) were detected in both species. Seroprevalence for MAP was significantly higher for domestic goats (26%) than for chamois (9.7%). In chamois, seroprevalence for pestivirus was higher in the west (6.5%) than in the east (range 0–1.8%), whereas seroprevalence for S. scabiei followed the opposite trend (west 4.6%; east 16.7–21.4%). We suggest that certain diseases could circulate between Cantabrian chamois and domestic goat populations, and domestic livestock may suppose a threat for the health status of sympatric Cantabrian chamois.     

Semen evaluation in the chamois of Abruzzi (Rupicapra pyrenaica ornata)

The chamois of Abruzzi (Rupicapra pyrenaica ornata) has been classified as endangered by the World Union for Conservation. The objective of this study was to analyze seasonal differences in the characteristics of various male reproductive organs and in semen quality. The study was conduced during 2004 in the reserve of Lama dei Peligni (Italy) on three chamois males aged between 2 and 5 years. Males were captured during March-May months and October-December months. Various testicular and scrotal measurements were taken and semen was collected using an electroejaculator. Sperm motility pattern was evaluated using computer assisted sperm analyzer, membrane integrity using differential staining and morphology with phase contrast microscopy. Testicular size, sperm motility membrane integrity and the percent of morphological normal spermatozoa were greater during October-December. The authors concluded that sperm characteristics are strongly influenced by season and that semen collected in this period (breeding season) has suitable quality for manipulation and long-term preservation.     

A panel of polymorphic microsatellites in the threatened Apennine chamois (Rupicapra pyrenaica ornata)

The Apennine chamois (Rupicapra pyrenaica ornata) is a highly endangered mountain ungulate. About 800 individuals still survive in central Apennines, Italy. Current conservation projects involve genetic identification of single animals, paternity tests, and assessment of relatedness. A suite of 60 microsatellite primer pairs from domestic bovids and cervids revealed polymorphisms for 10 loci, providing evidence for a low level of genetic variability, which might be a consequence of severe population bottlenecks and prolonged permanence at low numbers.     

Clinical infection of Cantabrian chamois (Rupicapra pyrenaica parva) by louping ill virus: new concern for mountain ungulate conservation?

Louping ill virus (LIV) was recently involved in an outbreak of encephalitis in domestic goats from Asturias region, northwestern Spain. Since livestock and wildlife in Asturias are frequently in close contact, we designed a retrospective survey for LIV antibody prevalence in wild ungulates by testing sera from 51 red deer (Cervus elaphus), 19 Cantabrian chamois (Rupicapra pyrenaica parva) and 8 roe deer (Capreolus capreolus) by the haemagglutination inhibition (HI) test. Only two Cantabrian chamois out of the 78 tested (2.6?±?3.5 %) gave positive results. Seroprevalence in chamois was 10.5?±?13.8 %. One of these chamois was found dead after falling down a cliff and the other one was found alive but with neurological signs. Histological examination of brain samples revealed that both animals showed severe inflammatory lesions compatible with a viral encephalitis caused by LIV, but LIV antigen was not detectable by specific immunohistochemistry. Real time RT-PCR was performed on formalin-fixed paraffin embedded sections of brain but was unable to confirm the presence of LIV RNA due to poor sample quality. By testing one of two HI positive sera from chamois by virus neutralization test and plaque reduction neutralization test against West Nile virus, Bagaza virus, Usutu virus, LIV and tick-borne encephalitis virus, we confirmed the presence of high antibody titres (1:10240) against LIV in the absence of antibodies to another Flavivirus. This work describes the first association between LIV and clinical encephalitis in chamois, which suggests that special attention should be paid to the impact on chamois conservation and management in Asturias, and perhaps in other European regions.     

Nunatak survival vs. tabula rasa in the Central Pyrenees: a study on the endemic plant species Borderea pyrenaica (Dioscoreaceae)

Aim  Borderea pyrenaica (Dioscoreaceae) is a Tertiary relict plant endemic to the Central Pyrenees. Because of its narrow distribution in a small geographical area and the fact that it is restricted to high alpine habitats, it constitutes an ideal model species for inferring the historical dynamics of population survival and migration during and after Quaternary glaciations in the Pyrenees.
Location  Central Pyrenees and pre-Pyrenees, Spain–France.
Methods  Eleven primer pairs were used to amplify 18 microsatellite loci in this allotetraploid species in a sample of 804 individuals from 15 populations, revealing a total of 77 alleles. Genotypic data of individuals and populations were analysed using clustering and Bayesian methods of analysis of population structure.
Results  A higher number of private alleles and a significantly higher allelic richness ( A *) were found in the southern area (21, A * = 2.295) than in the northern area (5, A * = 1.791). Furthermore, the allelic composition of the northern area represented a subset of that from the southern area.
Main conclusions  The hypothesis of in situ survival in northern Pyrenean nunataks was rejected, while peripheral refugia were considered to be restricted to the southern Pyrenees and pre-Pyrenees, where historical geographical fragmentation probably caused the divergence among southern Pyrenean populations. Molecular evidence indicates that these refugial populations probably colonized the northern area after sheet-ice retreat. Borderea pyrenaica lineages followed two migratory pathways in their northward colonization, suggesting several founder events for the populations that eventually reached the territory of the Gavarnie cirque.     

Temporal pooling of point transect data increases precision in density estimates of southern chamois

Estimating animal abundances in small areas is a difficult task and because a limited number of observations often results in low-precision estimates whose inaccuracies may even be exacerbated if surveys are focussed on clustered populations and/or are only carried out once a year. In an attempt to overcome this problem, we used point transects to monthly survey two small areas of a game reserve to assess the density of Pyrenean chamois (Rupicapra p. pyrenaica). The coefficient of variation associated with the density estimates after pooling observations by season was still high but decreased to reasonable values (<20%) when observations were over 29 chamois groups (clusters). Our results suggest that Distance Sampling may be a useful way of estimating the population density of mountain ungulates such as Pyrenean chamois in small rugged areas where only a small or moderate number of observations are to be expected.     

Late Pleistocene and Holocene chamois in Italy

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Characterization of seven (CTT)(n) microsatellite loci in the Pyrenean endemic Borderea pyrenaica (Dioscoreaceae): remarks on ploidy level and hybrid origin assessed through allozymes and microsatellite analyses

We present the identification and characterization of microsatellite loci in the Pyrenean endemic Borderea pyrenaica Miégeville (Dioscoreaceae). Seven microsatellite loci were isolated from a (CTT)(n)-enriched partial genomic library. Electropherograms patterns suggest that B. pyrenaica is a tetraploid species, as is its congener B. chouardii. One microsatellite locus was monomorphic, whereas the remaining ones presented from 2 to 10 alleles when analyzed in a sample of 60 individuals. Microsatellites have revealed higher levels of genetic variability than those in previous studies based on allozymes. Levels of genetic diversity are discussed in terms of tetrasomic (autotetraploidy) or duplicated disomic (allotetraploidy) modes of allele segregation. According to the first hypothesis, mean levels of genetic variability (H(min)-H(max)) range between 0.36 and 0.41, whereas, according to the second hypothesis, the 7 primer pairs amplified 11 chromosomal loci, and mean levels of observed and expected heterozygosities were 0.217 and 0.229, respectively, and did not differ significantly from HW expectations. These results suggest a hybrid allopolyploid origin for the Borderea taxa.     

Dynamics of an Infectious Keratoconjunctivitis Outbreak by Mycoplasma conjunctivae on Pyrenean Chamois Rupicapra p. pyrenaica

Between 2006 and 2008, an outbreak of Infectious Keratoconjunctivitis (IKC) affected Pyrenean chamois Rupicapra p. pyrenaica, an endemic subspecies of mountain ungulate that lives in the Pyrenees. The study focused on 14 mountain massifs (180,000 ha) where the species’ population is stable. Cases of IKC were detected in ten of the massifs and, in five of them, mortality was substantial. The outbreak spread quickly from the first location detected, with two peaks in mortality that affected one (2007) and three (2008) massifs. In the latter, the peak was seasonal (spring to autumn) and, in the former, the outbreak persisted through winter. To identify the outbreak’s aetiology, we examined 105 Pyrenean chamois clinically affected with IKC. TaqMan rt-PCR identified Mycoplasma conjunctivae in 93 (88.5%) of the chamois. Another rt-PCR detected Chlamydophila spp. in 14 of chamois, and 12 of those had mixed infections with mycoplasmas. In the period 2000–2007, the chamois population increased slightly (λ 1.026) but decreased significantly during the IKC outbreak (λ 0.8, 2007–2008; λ 0.85, 2008–2009) before increasing significantly after the outbreak (λ 1.1, 2009–2010). Sex-biased mortality shifted the adult sex ratio toward males (from 0.6 to 0.7 males per female) and reduced productivity slightly. Hunting was practically banned in the massifs where chamois experienced significant mortality and allowed again after the outbreak ended. Long-term monitoring of wild populations provides a basis for understanding the impacts of disease outbreaks and improves management decisions, particularly when species are subject to extractive exploitation.     

The successful introduction of the alpine marmot Marmota marmota in the Pyrenees,Iberian Peninsula,Western Europe

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Gene flow and hybridisation in a mixed oak forest (Quercus pyrenaica Willd. and Quercus petraea (Matts.) Liebl.) in central Spain

Oaks are long-standing models for the study of gene flow and hybridisation. Temperate (Quercus petraea) and sub-Mediterranean (Quercus pyrenaica) oaks coexist in central Spain, showing remarkable differences in population size and structure. Q. petraea has a scattered distribution in central Spain, where it is at one of the southernmost limits of its range, and forms low-density stands; in contrast, Q. pyrenaica is widespread in the region. We selected a mixed population of the two species ( approximately 13 ha, 176 adults and 96 saplings) to compare the patterns of gene flow within each species and the extent of introgression between them. Using five nuclear microsatellite markers, we performed a parentage analysis and found considerable immigration from outside the stand ( approximately 38% for Q. petraea and approximately 34% for Q. pyrenaica), and estimated average seed-dispersal distances of 42 and 14 m for Q. petraea and Q. pyrenaica, respectively. Introgression between species was also estimated using our microsatellite battery. First, we developed a multivariate discriminant approach and, second, we compared our results with a widely used clustering method (STRUCTURE). Both analyses were consistent with a low level of introgression between Q. petraea and Q. pyrenaica. Indeed, only 15 adult trees, approximately 8.5%, were identified as putative hybrids when both methods of analysis were combined. Hybrids may be most common in contact zones due merely to physical proximity.     

Recombination and the origin of sequence diversity in the DRB MHC class II locus in chamois (Rupicapra spp.)

We examined the evolutionary processes contributing to genetic diversity at the major histocompatibility complex (MHC) class II DRB locus in chamois (Rupicapra spp., subfamily Caprinae). We characterised the pattern of intragenic recombination (or homologous gene conversion) and quantified the amount of recombination in the genealogical history of the two chamois species, Pyrenean chamois (Rupicapra pyrenaica) and Alpine chamois (Rupicapra rupicapra). We found evidence for intragenic recombination, and the estimated amount of population recombination suggests that recombination has been a significant process in generating DRB allelic diversity in the genealogical history of the genus Rupicapra. Moreover, positive selection appears to act on the same peptide-binding residues in both analysed chamois species, but not in identical intensity. Recombination coupled with positive selection drives the rapid evolution at the peptide-binding sites in the MHC class II DRB gene. Many chamois MHC class II DRB alleles are thus much younger than previously assumed.