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.     

Hybridization Between Canada Lynx and Bobcats: Genetic Results and Management Implications

Hybridization between taxonomically similar species is an often-overlooked mechanism limiting the recovery of threatened and endangered species. We present molecular genetic data for the first time demonstrating that Canada lynx and bobcats hybridize in the wild. We verify that two microsatellite loci Lc106 and Lc110 have non-overlapping allele ranges between Canada lynx and bobcats, and that three putative lynx from Minnesota contain DNA from both bobcats and lynx. Additionally, we use a published test for the 16S rRNA region of mitochondrial DNA (mtDNA) to determine the maternal species; all hybrids had lynx mothers. Fifteen per cent (3/20) of our putative lynx samples were hybrids, although these data are not from a representative sampling effort. Hybridization may be an under-appreciated factor limiting the distribution and recovery of lynx. The presence of hybrids is thus a new factor in the population management of both species with potential implications for hunting and trapping of bobcats.     

Feeding ecology of the Iberian lynxLynx pardinus in eastern Sierra Morena (Southern Spain)

The feeding ecol ogy of the critically endangered Iberian lynxLynx pardinus Temminck, 1827, was studied in the most important remnant population, located in eastern Sierra Morena (Sierra de Andujar Natural Park, Spain). Three hundred sixty scats were collected from October 2001 through Sep tember 2002. RabbitsOryctolagus cuniculus dominated the lynx diet, as showed by frequency of occurrence (94%) and percent age of volume (91%). Red-legged partridgesAlectoris rufa were the most important secondary prey. No significant seasonal variation was observed regarding the diet preferences of the lynx. No difference was found in lynx diet by comparing two areas with high and low rabbit availability. Juvenile rabbits were positively selected when they were available, during late winter, spring and early summer.     

Diet of Eurasian lynx <Emphasis Type="Italic">Lynx lynx</Emphasis> in the northern Dinaric Mountains (Slovenia and Croatia)

The Eurasian lynx Lynx lynx (Linnaeus 1758) is an opportunistic predator that usually selects the smallest ungulate available. Its diet varies considerably among different regions; therefore it is important to study lynx diet in different parts of the species’ range. We studied the diet of lynx from the endangered Dinaric population in Slovenia and Croatia by analyzing lynx scats, prey remains, and stomach contents. Dinaric lynx mainly killed European roe deer Capreolus capreolus (0.64 frequency of occurrence, 79% of all consumed biomass), which were used more frequently during winter and spring. Ungulates were killed more often by adult males than by lynx of other age and sex groups. In contrast to studies from other regions, lynx in the northern Dinaric Mountains also frequently fed on the edible dormice Glis glis (0.18 frequency of occurrence, 7% of all consumed biomass). This large rodent appears to be an important alternative prey, especially for females and young lynx, and was the reason for the highest use of rodents reported so far for the Eurasian lynx. Edible dormice in Dinaric forests have highly variable numbers of active animals. Seasonal and possibly annual variation in dormouse availability obviously affects lynx diet. This is a rare example where variability in the availability of the alternative prey and not the preferred prey leads to the dietary shift. This study confirms the opportunistic nature of Eurasian lynx and the regional variability of its diet.     

Detection of extended-spectrum beta-lactamase-producing Escherichia coli isolates in faecal samples of Iberian lynx

Aims: To characterize the diversity of extended‐spectrum beta‐lactamase (ESBL)‐producing Escherichia coli isolates recovered within the faecal microbiota of Iberian lynx. The identification of other associated resistance genes and the analysis of clonal relationship were also focused in this study. Methods and Results: From 2008 to 2010, 128 faecal samples of Iberian lynx (wild and captive animals) were collected. Eleven tested samples contained cefotaxime‐resistant E. coli isolates (all belonging to captive animals) and 10 ESBL‐producing isolates were showed. CTX‐M‐14 and SHV‐12 ESBL‐types were detected and seven different patterns were identified by pulsed‐field gel electrophoresis analysis. Conclusions: The occurrence of unrelated multiresistant E. coli in faecal flora of captive specimens of Iberian lynx, including the presence of ESBLs, resistant genes in integrons and virulence determinants was showed in this study. Significance and Impact of the Study: The results obtained in this study highlight the environmental problem as future reintroductions of Iberian lynx could lead to a spread of resistant bacteria. Additionally, ESBL‐producing bacteria can represent a health problem for this endangered species.     

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.     

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   

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     

To Eat or Not To Eat? The Diet of the Endangered Iberian Wolf (Canis lupus signatus) in a Human-Dominated Landscape in Central Portugal

Livestock predation by large carnivores and their persecution by local communities are major conservation concerns. In order to prevent speculations and reduce conflicts, it is crucial to get detailed and accurate data on predators’ dietary ecology, which is particularly important in human dominated landscapes where livestock densities are high. This is the case of the endangered Iberian wolf in Portugal, an endemic subspecies of the Iberian Peninsula, which has seen its population distribution and abundance decline throughout the 20th century. Accordingly, the diet of the Iberian wolf was analyzed, using scat analysis, in a humanized landscape in central Portugal. From 2011 to 2014, a total of 295 wolf scats were collected from transects distributed throughout the study area, prospected on a monthly basis. Scat analysis indicated a high dependence of Iberian wolf on livestock. Domestic goat predominated the diet (62% of the scats), followed by cow (20%) and sheep (13%); the only wild ungulate present in the scat analysis was the wild boar (4% of the scats). Our results show that even though livestock constitute most part of wolves diet, different livestock species may represent different predation opportunities. We conclude that the high levels of livestock consumption may be a result of low diversity and density of wild ungulates that settles livestock as the only abundant prey for wolves. Our findings help on the understanding of the Iberian wolf feeding ecology and have implications for conflict management strategies. Finally, management implications are discussed and solutions are recommended.     

Factors conditioning the camera-trapping efficiency for the Iberian lynx (Lynx pardinus)

Camera trapping is the most used method for surveying medium-sized carnivores in Spain. The main target for these surveys has been the Iberian lynx, the most endangered cat in the world. The Iberian lynx conservation program has received the largest EU LIFE projects grant. So, efficiency is a key goal for managing this grant. During 2003 and 2007, we have applied these funds to the survey of the Iberian lynx in Eastern Sierra Morena (Spain). Using two different techniques, we have studied both to see which is the most efficient. The survey developed in active latrines resulted more efficient than that of scent stations and live prey camera trapping throughout the years, although there has been a variation between years. Otherwise, the live prey method has been the one providing the greatest speed and number of pictures per entrance. We suggest that camera-trapping surveys can be improved in terms of efficiency for a wide range of species, or at least for the Iberian lynx. To improve the results, cameras might be placed in relation to breeding territories. With this determinant, camera-trapping surveys would be shorter than 120 days. Finally, we suggest how those surveys for medium carnivores should be designed.     

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.     

Evaluation of Camera Trapping for Estimating Red Fox Abundance

ABSTRACT The nature reserve Serra da Malcata, Portugal, was recently considered a site for Iberian lynx (Lynx pardinus) reintroduction. Because of potential disease risk posed by red foxes (Vulpes vulpes) in the area, a reliable estimate of fox abundance was critical for a dependable reintroduction program. We adapted camera-trapping techniques for estimating red fox abundance in the reserve. From July 2005 to August 2007, we conducted 7 camera-trapping sessions, allowing for individual identification of foxes by physical characteristics. We estimated abundance using the heterogeneity (M_h) model of the software program CAPTURE. Estimated density ranged from 0.91 ± 0.12 foxes/km²to 0.74 ± 0.02 foxes/km². By estimating red fox density, it is possible to define the number of foxes that must be sampled to assess the presence of potential fox-transmitted diseases that may affect lynx reintroduction.     

Diet of the Iberian hare (Lepus granatensis) in a mountain ecosystem

The diet of the Iberian hare (Lepus granatensis) was studied through microhistological pellet analysis in two areas from a mountain ecosystem in Central Portugal. Fecal pellets were collected monthly in 24 plots spatially distributed throughout the two study areas. For each period, a sample of 15 to 20 pellets was milled and 400 epidermal fragments were identified, by comparison with a reference collection. A wide range of plant species was observed in hare’s diet. Grasses represent the basis of the Iberian hare diet, with frequencies always higher than 50% in both study areas (annual average = 69.98%). Most of the 35 species of grasses assembled for the reference collection (91.43%) were identified in the pellets. Nevertheless, only six of these were consumed in proportions greater than 5%, being Anthoxanthum odoratum, Secale cereale and Agrostis spp. the species ingested in higher frequencies. The rate of grasses consumption reached 80.69% in winter but decreased in summer to around 55%. In this season, a concurrent rise in the ingestion of other plant groups, like herbs and shrubs, and of plant inflorescences was observed. This work provides the first results on the Iberian hare’s diet on mountain ecosystems, and suggests that the Iberian hare diet in a mountain ecosystem is similar to the observed in L. europaeus and L. timidus.     

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.     

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.     

Using historical accounts to set conservation baselines: the case of Lynx species in Spain

The knowledge of the historical range of organisms is necessary to understand distribution dynamics and their drivers as well as to set reference conditions and conservation goals. We reviewed written sources documenting the presence of lynxes in Spain between the sixteenth and nineteenth centuries, trying to infer whether Lynx records referred to the Iberian (Lynx pardinus) or the Eurasian (Lynx lynx) species. We compiled 151 spatially specific, non-redundant Lynx records, dating between 1572 and 1897. Records appeared split in two neatly separated areas, North and South. Arguably, all historical records from the South area correspond to Iberian lynx, since almost all of them fall within the estimated range of the species in the 1950s. This suggests that the core distribution area of the species remained relatively stable for centuries, until the second half of the twentieth century. The reported range in the 1950s is thus an appropriate scenario to set conservation goals for this critically endangered species. The North area extended from Galicia and northern Portugal to the Mediterranean, mainly occupying the Atlantic-climate area of northern Iberia. Reported direct measurements and the accumulation of indirect evidences suggest that most, if not all, northern Lynx records refer to the Eurasian lynx, which had been previously detected in archaeological sites of the area. This species was apparently present in the Iberian Peninsula until the early-nineteenth century. Our results provide an objective, baseline distribution of Lynx species in Spain useful to guide conservation efforts both at the Iberian and European levels.     

Mesopredator behavioral response to olfactory signals of an apex predator

Olfactory signals constitute an important mechanism in interspecific interactions, but little is known regarding their role in communication between predator species. We analyzed the behavioral responses of a mesopredator, the red fox (Vulpes vulpes), to an olfactory cue (scat) of an apex predator, the lynx (Lynx lynx) in Bia?owie?a Primeval Forest, Poland, using video camera traps. Red fox visited sites with scats more often than expected and the duration of their visits was longer at scat sites than at control sites (no scat added). Vigilant behavior, sniffing and scent marking (including over-marking) occurred more often at scat sites compared to control sites, where foxes mainly passed by. Vigilance was most pronounced during the first days of the recordings. Red fox behavior was also influenced by foxes previously visiting scat sites. They sniffed and scent marked (multiple over-marking) more frequently when the lynx scat had been over-marked previously by red fox. Fox visits to lynx scats may be seen as a trade-off between obtaining information on a potential food source (prey killed by lynx) and the potential risk of predation by an apex predator.     

Space use and habitat preferences of the invasive American mink (<Emphasis Type="Italic">Mustela vison</Emphasis>) in a Mediterranean area

Space use, intra-territorial habitat preferences, and factors affecting both were studied in an invading population of American mink, Mustela vison, in two rivers of a Mediterranean region of Spain. Average linear home range was 1.19 ± 0.73 km (±SD) and core area was 0.21 ± 0.08 km for resident males (n = 10); while for females (n = 5) they were 0.54 ± 0.14 and 0.19 ± 0.11 km, respectively. Overlapping between the home ranges of residents was low. In no case their core areas overlapped. Home ranges were small in comparison to other study areas and in general the resident minks were territorial. Linear home range length was related to individual weight and to the river. Weight had a positive effect indicating a potential body condition effect, while river may be showing a habitat quality effect. Habitat preferences were positively affected by the abundance of helophytic vegetation and negatively by the presence of human activity. Helophytic vegetation offers both food and refuges, while human activity may represent a potential danger. Percentage of captures was higher inside the core areas and was slightly influenced positively by abundance of helophytic vegetation. All this information should be considered when designing and implementing measures to control the expansion of American minks. We recommend keeping going with the trapping sessions but, given the results obtained, reducing the distance between traps down to 200 m to maximize capturability (i.e., about doubling the trapping effort), and, when available, placing them near helophytic vegetation. In the absence of helophytic vegetation, traps should be located near any kind of vegetation providing coverage for mink and far from human activity.     

Feeding responses of the red fox (<Emphasis Type="Italic">Vulpes vulpes</Emphasis>) to different wild rabbit (<Emphasis Type="Italic">Oryctolagus cuniculus</Emphasis>) densities: a regional approach

We investigate the feeding responses of the red fox (Vulpes vulpes) at a regional scale to different densities of European wild rabbit (Oryctolagus cuniculus) in central–southern Spain. Rabbit abundance indices were obtained in 86 localities during summer 2002. The diet of the fox was studied by analysis of 114 scats collected in 47 of these localities. The feeding response of the fox was examined by a representation of the dry weight percent of rabbit in the diet as a function of the abundance of rabbits; this used data only from those localities where at least 3 scats were collected (70 fox scats from 18 localities). We evaluated the relationship between rabbit abundance and the diversity of the diet of the fox. The feeding patterns of red foxes approximated to Holling’s type III functional response, typical of opportunistic predators. There was a negative relationship between the diversity of the fox’s diet and the abundance of rabbits. Therefore, the fox apparently behaves as a facultative predator, feeding on rabbits when they are abundant and shifting to other prey (and hence a more diverse diet) when rabbits are scarce. These findings are the first step towards understanding the potential role of red foxes in regulating rabbit populations in central–southern Spain.     

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.