Phylogenetic and phylogeographic analysis of Iberian lynx populations

The Iberian lynx (Lynx pardinus), one of the world's most endangered cat species, is vulnerable due to habitat loss, increased fragmentation of populations, and precipitous demographic reductions. An understanding of Iberian lynx evolutionary history is necessary to develop rational management plans for the species. Our objectives were to assess Iberian lynx genetic diversity at three evolutionary timescales. First we analyzed mitochondrial DNA (mtDNA) sequence variation to position the Iberian lynx relative to other species of the genus LYNX: We then assessed the pattern of mtDNA variation of isolated populations across the Iberian Peninsula. Finally we estimated levels of gene flow between two of the most important remaining lynx populations (Do?ana National Park and the Sierra Morena Mountains) and characterized the extent of microsatellite locus variation in these populations. Phylogenetic analyses of 1613 bp of mtDNA sequence variation supports the hypothesis that the Iberian lynx, Eurasian lynx, and Canadian lynx diverged within a short time period around 1.53-1.68 million years ago, and that the Iberian lynx and Eurasian lynx are sister taxa. Relative to most other felid species, genetic variation in mtDNA genes and nuclear microsatellites were reduced in Iberian lynx, suggesting that they experienced a fairly severe demographic bottleneck. In addition, the effects of more recent reductions in gene flow and population size are being manifested in local patterns of molecular genetic variation. These data, combined with recent studies modeling the viability of Iberian lynx populations, should provide greater urgency for the development and implementation of rational in situ and ex situ conservation plans.     

Estimation of the Iberian lynx (Lynx pardinus) population in the Do?ana area, SW Spain, using capture�Crecapture analysis of camera-trapping data

The Iberian lynx (Lynx pardinus) has a highly restricted geographic distribution, limited even within the Iberian Peninsula. The last national survey reported less than 200 remaining individuals, distributed in two isolated areas??Andújar-Cardeña and Doñana??and in consequence, the Iberian lynx was listed by the International Union for Conservation of Nature as ??Critically Endangered??. In this study, we estimate the Iberian lynx population size in the Doñana area using capture?Crecapture analysis of camera-trapping data. A model with different capture probability for each individual (M_h) yielded an estimate of 26 Iberian lynxes (SE?=?5.26) more than 1 year old. It is considered that a small slant in the estimation of the number of individuals could exist due to the presence of dispersers inside the study area that were not detected. Our study shows: (1) a reduction in number since the 1980s (45 individuals), and falling below the theoretical threshold of genetic viability, (2) changes in the species?? spatial distribution in this area, and (3) as for other carnivore species, photographic capture?Crecapture methods are applicable for estimating the size of Iberian lynx populations     

Historical DNA reveals the phylogenetic position of the extinct Alpine lynx

During the last two centuries, lynx populations have undergone severe declines and extinctions in Europe. The Alpine lynx, once distributed across the whole Alpine arc, became extinct due to direct human prosecution and deprivation of its main prey in the 1930s. Similar to the Iberian lynx Lynx pardinus , its taxonomy has been subject to several controversies. Moreover, knowing the taxonomic status of the Alpine lynx will help to define conservation units of extant lynx populations in Europe. In this study, we investigated two mitochondrial DNA regions in museum specimens ( n =15) representing the autochthonous Alpine population and in samples from extant Eurasian lynx Lynx lynx populations in Europe and Asia ( n =17). Phylogenetic analysis (cytochrome b , 345 bp) placed the Alpine lynx within the Eurasian lynx lineage. Among all individuals examined, seven different haplotypes (control region, 300 bp) were observed but no unique Alpine haplotype was discovered. Haplotypes of the extinct Alpine population were identical to previously described haplotypes in Scandinavian lynx signifying a recent genetic ancestry with current European populations. Moreover, our genetic data suggest two distinct glacial refugia for the Carpathian and Balkan population. Overall this study demonstrates that historical DNA from extinct populations can help to disentangle the phylogenetic relationships and historical biogeography of taxa with only a limited number of extant populations remaining.     

Faecal genetic analysis to determine the presence and distribution of elusive carnivores: design and feasibility for the Iberian lynx

Noninvasive methods using genetic markers have been suggested as ways to overcome difficulties associated with documenting the presence of elusive species. We present and assess a novel, reliable and effective molecular genetic technique for the unequivocal genetic identification of faeces from the endangered Iberian lynx (Lynx pardinus). From mitochondrial DNA (mtDNA) cytochrome b and D-loop region sequences, we designed four species-specific primers (for products 130-161 bp long) that were considered to be likely to amplify degraded DNA. We compared two DNA extraction methods, various DNA amplification conditions and the robustness and specificity of the primer pairs with 87 lynx samples from 5 potentially different lynx populations and with 328 samples of other carnivore species. The utility of the identification technique was tested with faeces of different ages, with faeces from controlled field experiments, and with faeces collected from locales with possible lynx populations from throughout the state of Andalusia, Spain (8052 km2). Faecal mtDNA extraction was more efficient using PBS wash of the faeces instead of a faeces homogenate. Our assay increased from 92.6 to 99% efficiency with a second amplification and a reduction in template concentration to overcome polymerase chain reaction (PCR) inhibition. Our assay never produced false positives, and correctly identified all lynx faeces. Of 252 faeces samples of unknown species collected throughout Andalusia, 26.6% (from three different areas) were classified as Iberian lynx, 1.4% showed evidence of PCR inhibition and 1.2% were of uncertain origin. This method has proven to be a reliable technique that can be incorporated into large-scale surveys of Iberian lynx populations and exemplifies an approach that can easily be extended to other species.     

Genetics at the verge of extinction: insights from the Iberian lynx

Population viability might become compromised by the loss of genetic diversity and the accumulation of inbreeding resulting from population decline and fragmentation. The Iberian lynx (Lynx pardinus) provides a paradigmatic example of a species at the verge of extinction, and because of the well‐documented and different demographic histories of the two remaining populations (Doñana and Andújar), it provides the opportunity to evaluate the performance of analytical methods commonly applied to recently declined populations. We used mitochondrial sequences and 36 microsatellite markers to evaluate the current genetic status of the species and to assess the genetic signatures of its past history. Mitochondrial diversity was extremely low with only two haplotypes, alternatively fixed in each population. Both remnant populations have low levels of genetic diversity at microsatellite markers, particularly the population from Doñana, and genetic differentiation between the two populations is high. Bayesian coalescent‐based methods suggest an earlier decline starting hundreds of years ago, while heterozygosity excess and M‐ratio tests did not provide conclusive and consistent evidence for recent bottlenecks. Also, a model of gene flow received overwhelming support over a model of pure drift. Results that are in conflict with the known recent demography of the species call for caution in the use of these methods, especially when no information on previous demographic history is available. Overall, our results suggest that current genetic patterns in the Iberian lynx are mainly the result of its recent decline and fragmentation and alerts on possible genetic risks for its persistence. Conservation strategies should explicitly consider this threat and incorporate an integrated genetic management of wild, captive and re‐introduced populations, including genetic restoration through translocations.     

Effects of food supplementation on home-range size, reproductive success, productivity and recruitment in a small population of Iberian lynx

In a conservation context, food supplementation is a management tool used to reverse the decline of food-limited populations by means of positive changes in behaviour and fitness that may be reflected in population parameters. The critically endangered Iberian lynx Lynx pardinus has suffered a dramatic decline primarily because of the severe drop of its main prey, the European wild rabbit Oryctolagus cuniculus . To reverse this situation, a food supplementation programme has been implemented in Doñana, south-west Spain, since 2002. In this study, we assess the utility of providing artificial food to reduce home-range (HR) size, and to increase productivity, survival and recruitment in a scenario of low lynx density, as compared with reference data from the same population in the absence of extra food. Food supplementation produced a significant contraction of core areas, but not of complete lynx HRs. We did not detect any significant change in productivity or dispersal rates, but supplementation could have helped transient adult lynx to settle down. The positive effects of food supplementation may have been partly countered by factors such as inbreeding, Allee effects and disease outbreaks, whose effects may have been exacerbated in this small lynx population. Food supplementation, however, proved useful to retain individuals, to keep range sizes within their normal range of values, thus maintaining spatial organization, and to allow lynx reproduction and kitten survival in areas with very low prey density. Therefore, we recommend keeping an extensive and intensive supplementary feeding programme until the density of wild rabbits will enable the viability of this endangered lynx population.     

Conservation biogeography of ecologically interacting species: the case of the Iberian lynx and the European rabbit

Aim To relate the recent Iberian lynx decline to changes in the distribution of the European rabbit after the haemorrhagic disease outbreak of 1989. As Iberian rabbits evolved in two geographically separated lineages, being the recent lynx range practically restricted to the southwestern lineage, we also test if differential range dynamics exists for these lineages, with the consequent implications for lynx conservation and reintroduction planning. Location The Iberian Peninsula. Methods We modelled environmental favourability for the lynx based on its distribution before 1989, and for the rabbit using distribution data collected primarily after 1989, and validated them using independent abundance data. We compared both models and combined them in a lynx occurrence forecast. We correlated the prevalence of southwestern rabbit lineage with the environmental favourability for the rabbit. Results The environmental lynx model correlated with past lynx abundance data, but did not reflect its recent strong range contraction. The rabbit model correlated with recent rabbit abundance, but was negatively correlated with the environmental model for the lynx. The combination of both models forecasted lynx occurrence in a few separated nuclei, which encompass all recent lynx records. The prevalence of rabbit's southwestern lineage correlated negatively with favourability for the rabbit. Main conclusions The region to which the lynx became confined before 1989 is currently less favourable for rabbits, whereas more favourable areas remain outside lynx reach. This differential favourability correlates with rabbit phylogeographical structure, suggesting that the southwestern lineage is facing more unfavourable conditions or is less resilient to recent diseases. The loss of concordance between lynx distribution and the whole rabbit phylogeographical structure has prevented lynx persistence in northeastern rabbit lineage areas, which should be considered in lynx reintroduction planning. Similar conservation problems could affect other ecologically interacting species whose distributions’ overlapping has sharply diminished.     

Looking for the Iberian lynx in central Spain: a needle in a haystack?

The Iberian lynx Lynx pardinus has suffered a dramatic reduction in its range throughout the Iberian Peninsula and at present is one of the most endangered mammals in the world. The latest studies report that, out of the 48 breeding areas that existed in 1990, only two populations are left in southern Spain. As a consequence, some of the formerly largest populations, such as Montes de Toledo (central Spain), are to all intents and purposes regarded as extinct. To determine the current distribution of Iberian lynx outside the two recognized populations, we surveyed five different areas where the species is considered extinct and collected 581 faeces for the genetic identification of the species. We identified 18 samples as belonging to Iberian lynx in four out of the five areas studied, providing clear evidence for the presence of lynx in central Spain. In some areas the species was detected repeatedly at different localities and on different dates, indicating a regular occurrence of an unknown number of individuals. The conservation implications of these results are discussed in terms of the genetic importance of the individuals found and future reintroductions of the species from an ongoing captive-breeding programme.     

Human–felid conflict as a further handicap to the conservation of the critically endangered Iberian lynx

Most studies on felid depredation of livestock have focused on big cats, and little attention has been paid to this type of conflict in smaller species. The medium-sized Iberian lynx (Lynx pardinus) is not thought to be affected by conflict with humans. However, parallel to an increase in the range of the Iberian lynx in Andújar-Cardeña, an increased incidence of Iberian lynx attacks on livestock has been recorded. A 6-year overview of Iberian lynx predation on livestock in this population shows a total of 40 attacks involving 716 kills (31 attacks on poultry and nine on sheep). Although the majority of these attacks (78 %) were carried out against poultry, sheep depredation resulted in higher economic losses, mainly in extensive flocks (4.6 times more than semi-intensive flocks). An effective compensation program has been implemented in order to mitigate the consequences of the human–lynx conflict in this area. Given that this sort of conflict could become an acute impediment to future conservation of the most endangered felid, managers should anticipate and prevent the potential conflicts that could arise as Iberian lynx colonizes more developed areas.     

A comparison of scat counts and camera-trapping as means of assessing Iberian lynx abundance

Rigorous cost-effective census methods are an essential part of managing wild animal populations, particularly if the involved species require continuous monitoring. In recent years, the range and size of the Iberian lynx (Lynx pardinus) population have increased notably as a result of conservation programs. Given this expansive scenario, monitoring Iberian lynx using camera-traps may become logistically and financially unfeasible. Here, we test whether or not scat counts are as efficient as camera-trapping in predicting lynx densities and compare the cost of these two approaches to lynx monitoring. We found a significant positive linear relationship between these two methods of assessing Iberian lynx abundance. Lynx abundance assessment by scat counting was six times less expensive than camera-trapping; there was no significant effect of the two study areas on the observed patterns (GLM F_1,7?=?45.81, P?相似文献   

Late Pleistocene and Holocene development of the felid fauna (Felidae) of Europe: a review

In the fossil deposits of the Late Pleistocene ( c . 115 000–11 500 years ago), five felid species are recorded in Europe: the wildcat Felis silvestris , Eurasian lynx Lynx lynx , Iberian lynx Lynx pardinus , leopard Panthera pardus and cave lion Panthera leo spelaea . In the Holocene, Europe was colonized by F. silvestris , L. lynx and L. pardinus as well as the lion Panthera leo . The status of P. pardus in post-glacial Europe is unknown. So far, only sparse records indicate that P. pardus survived into the early Holocene. During the Late Glacial, both L. lynx and L. pardinus occurred on the Iberian Peninsula. However, from the Holocene, only the Iberian lynx is recorded in this region. There are subfossil records that indicate that L. pardinus also occurred in central and western France until c . 3000 years ago. Surprisingly, with reservations on the determination of the bones (by J. Altuna), both lions and cave lions seem to be recorded in the Iberian Peninsula in the Late Glacial. There are published records of the lion P. leo in the northern Iberian Peninsula from the early Holocene. However, its presence in Europe on the basis of subfossil records was proven initially from the Atlantic period. In Ponto-Mediterranean regions of Europe, the lion is recorded from the Atlantic to the younger sub-Atlantic.     

The importance of genetic variability and population differentiation in the Eurasian lynx Lynx lynx for conservation,in the context of habitat and climate change

• 1 The Eurasian lynx Lynx lynx occupies a variety of environmental and climatic conditions, and the majority of present‐day European populations have either recovered from severe demographic bottlenecks, or are living in fragmented habitat. These factors may have affected the genetic variability of lynx populations. We summarize available data on genetics, population status and ecology of these felids to shed light on the pattern and mechanisms behind their genetic variability and population differentiation in Europe.
• 2 Genetic studies conducted so far, based on mtDNA and microsatellites, have shown that the Eurasian lynx has low to moderate genetic variability. Variability is lowest in the north (Scandinavian bottlenecked population), but is also low in the Carpathian region. A trend towards loss of genetic variation has been noted in fragmented and reintroduced populations. Genetically, the populations are highly differentiated from each other.
• 3 There are clear relationships between the pattern of lynx genetic variability, differentiation between the populations, and such factors as population history (demographic bottlenecks), social interactions and habitat fragmentation. The genetic divergence between lynx populations is also strongly correlated with the depth and duration of snow cover.
• 4 Our review provides evidence that the lynx is undergoing significant genetic differentiation, due to several factors. To enable better planning of conservation programmes for the Eurasian lynx, researchers should identify the Evolutionarily Significant Units among its populations, using different classes of molecular markers.

     

Evaluating mortality rates and causalities in a critically endangered felid across its whole distribution range

The conservation of endangered species requires accurate data, and knowledge of cause-specific mortality rates is one of the most important issues. In recent years, conservation programs for the critically endangered Iberian lynx Lynx pardinus have been developed on the basis of mortality data derived 30 years ago from the small Doñana population. Thus, there is an urgent need for an update of mortality rates and causes in both populations (Sierra Morena and Doñana). Here we use radio-tracking information from the whole range of the Iberian lynx to quantify mortality rates and identify their causes. Between 2006 and 2011, we radio-tagged 78 Iberian lynxes from its two remaining populations (39 from Sierra Morena and 39 from Doñana). Mortality events were evaluated to identify causes, and cause-specific annual mortality rates (AMR) were obtained using the nonparametric cumulative incidence function estimator. Overall, AMR was estimated at 0.16?±?0.05 (0.19?±?0.09 in Sierra Morena and 0.12?±?0.07 in Doñana). Disease was the main cause of mortality both for the whole population and the Doñana population. Poaching was the main cause of mortality in Sierra Morena. Our results suggest that the best strategy for conserving this species is to focus action on decreasing the fatal effect of disease and poaching. Given the possible existence of an underlying inbreeding-mediated immunosuppression, genetic management aimed at increasing the genetic diversity of this population is also recommended.     

First report of an intraerythrocytic small piroplasm in wild Iberian lynx (Lynx pardinus)

A wild injured Iberian lynx (Lynx pardinus) was taken from the Sierra Morena population. During the health check small intraerythrocytic piroplasms, morphologically indistinguishable from other feline piroplasms, were observed in Wright-Giemsa-stained blood films. Amplification by polymerase chain reaction of a portion of the 18S nuclear small subunit (NSS) rRNA gene and sequencing revealed similarity of the unknown organism with sequences obtained from Pallas's cat from Mongolia and from a domestic cat in Spain. In a retrospective (1993-2003) study of 50 Iberian lynx tissue samples, no amplifications of the 18S NSS rRNA gene of the organism were obtained. This is the first report of a naturally occurring erythroparasitemia in the Iberian lynx and the first documented case of naturally occurring piroplasm infection in a free-ranging felid from Europe.     

Assessment of Canada Lynx Research and Conservation Needs in the Southern Range: Another Kick at the Cat

Abstract The ecology of Canada lynx (Lynx canadensis) and their main prey, snowshoe hares (Lepus americanus), is poorly understood in southern Canada and the contiguous United States compared to the boreal forest of Canada and Alaska, USA, where both species are well studied. However, given recent listing of lynx under the Endangered Species Act, accurate understanding of lynx and snowshoe hare ecology and conservation requirements in the United States is a high priority. We critically examined unchallenged perceptions and important research needs related to lynx and hare ecology and conservation at the southern extent of their range. Contrary to popular dogma, lynx do not require old-growth forest for denning, but further research on lynx and hare use of fragmented landscapes at lower latitudes is required. The contention that southern lynx are subject to higher interference or exploitative competition compared to their northern counterparts remains without strong empirical support. Lynx rely more on red squirrels (Tamiasciurus hudsonicus) and possibly other alternate prey at lower latitudes, but hares are the predominant food type for lynx across their range. Southern lynx and hare populations do not exhibit periodic cyclicity, but harvest statistics suggest that lynx abundance in the southern range is highly variable, implying that numerical fluctuations likely are fueled by immigration from Canada. Southern lynx population viability in the absence of ingress is suspect and thus maintaining connectivity with northern areas of occupancy should be a priority. Successful conservation of lynx populations in the contiguous United States will require 1) improved understanding of lynx population and habitat ecology at lower latitudes, 2) protection and management of large tracts of lynx and snowshoe hare habitat, and 3) ensured connectivity between lynx populations at the core and periphery of the species' range. However, in light of the numerous challenges facing conservation of populations of many species at their southern distributional limit, the long-term prognosis for lynx in the southern range currently is uncertain.     

First breeding record of a one-year-old female Iberian lynx

According to literature, female Iberian lynx (Lynx pardinus) start reproducing at 3 years of age. In more recent years, females breeding at 2 years have been recorded in wild populations. Here, however, we describe the first breeding record for a one-year-old female Iberian lynx. This finding was obtained by camera-trapping data, provided by the yearly monitorization of the species that is carried out across its distribution area. A litter of two lynx kittens was found at the south-east limit of the Andujar-Cardeña population. Two camera-traps were set up near the den to identify the breeding female. Photographs obtained confirmed that the breeding female had already been identified the previous year at less than 3 months of age. The case reported here of a yearling female breeding is the first ever from historical distribution range of the species, and as such, is an exceptional event in the reproduction of the Iberian lynx.     

Last lynxes in Portugal? Molecular approaches in a pre-extinction scenario

The Iberian lynx is the most threatened felidin the world and has suffered a declinethroughout its range. Effective monitoring ofthe species' presence is essential. Fieldwork inpreviously identified areas of lynx occurrencein Portugal has resulted in the collection of104 possible lynx scats. Recently, there hasbeen little or no evidence of lynx presence andscats could be confused with others from moreabundant carnivores such as wildcat, fox anddog. In order to confirm or not exclude thepresence of the species, identification ofscats was performed through the amplificationof lynx-specific mitochondrial DNA sequences.Two samples collected in Malcata NaturalReserve in 1997 were identified as lynx. Thisis the most recent and reliable proof of lynxpresence in Portugal^*. Given the territorialbehavior of lynx, stable resident populationswould have produced a higher proportion ofpositively identified scats. Local extinctionsmight have taken place, and this genetic datasupports a suspected national pre-extinctionscenario for the species. Genetic analysisusing a non-invasive approach has proved to bean informative part of the lynx monitoringprogram. Technical problems faced and overcomeare also presented.     

Distribution and status of lynx in the border region between Czech Republic, Germany and Austria

This paper summarizes available information concerning the presence of the Eurasian lynxLynx lynx Linneaus, 1758 in the Šumava Mountains and adjacent areas along the common border of Czech Republic, Germany and Austria. Our data give an overview of the lynx population occupying the border region between the three countries from 1990 to 1999. We estimated population size using radiotracking data. From 1990 to 1998, population increased from under 20 to nearly 70 resident animals. During this time, reproduction increased as well, with a maximum of 55 kittens observed in the rearing period of 1998 to 1999. Mortality data indicated that illegal hunting was widespread. Our paper discusses possible links with other lynx populations and describes the legal status of lynx in the three different countries. Current management approaches are outlined and steps toward a long-term conservation plan for the population are proposed.     

Epidemiology of Ancylostoma spp. in the endangered Iberian lynx (Lynx pardinus) in the Doñana National Park, south-west Spain

The epidemiology of Ancylostoma spp. was studied in the endangered Iberian lynx (Lynx pardinus) in the Do?ana National Park, south-west Spain. Faecal samples were collected throughout a complete annual cycle (August 1997 to September 1998). The overall egg prevalence of Ancylostoma spp. was 57.8%. The pattern of abundance of Ancylostoma spp. eggs in faeces was overdispersed. Juvenile lynx demonstrated a statistically higher prevalence and abundance of Ancylostoma spp. than in adults. These levels of egg output (maximum 21195 epg), as previously reported in free ranging large felid cubs, could be close to disease involvement. The potential pathogenicity of hookworms and the influence of individual and ecological factors on hookworm transmission in the Iberian lynx from the Do?ana National Park population are discussed.     

Genetic structure of the Eurasian lynx population in north-eastern Poland and the Baltic states

We analyzed the genotypes of Eurasian lynx (Lynx lynx) from three populations in the westernmost part of the species main range. One population was situated at the distribution edge (NE Poland) and the two other (Latvia and Estonia) were located within the main, contiguous range of the species. The aim was to determine if the genetic composition varied among these populations and if there was evidence of isolation among them. Based on microsatellite allele frequencies, we found the allelic richness in Polish lynx to be lower than that in lynx from Latvia and Estonia. We also found significant differentiation among the lynx populations, with the NE Poland population forming a distinct genetic group relative to the two other populations (R _ST = 0.15 and 0.22, P < 0.0001). We suggest that genetic differentiation among lynx populations is the result of habitat insularisation that limits gene flow. This finding emphasizes the necessity to consider the lynx genetic differentiation in conservation planning of this species in Poland.