Lepus Timidus Research Articles

I: ORIGINS AND ANCESTORS

The Irish Hares are an off-shoot of the much older Hare family in Norfolk. They go back to a certain Michael Hare ‘of Monktown in County Dublin’, who had come to Ireland from Norfolk in the 17th Century, and died without issue in 1685, leaving a large fortune in a Will dated 1684 to his nephew John Hare, ‘a native of Norfolk’. There is evidence of the relationship between the Irish and the Norfolk Hares in [the] similarity between the family coat of arms, particularly in their shields, which can be traced back by the Norfolk branch of the family to the Norman family of D'Harcourt which is believed to have fought in the Crusades.

Spatial and temporal variation in mountain hare (Lepus timidus) abundance in relation to red grouse (Lagopus lagopus scotica) management in Scotland

Mountain hares (Lepus timidus) are regularly harvested on Scottish moorland managed for recreational hunting of red grouse (Lagopus lagopus scotica). There are concerns that large-scale culls of mountain hares on grouse moors are contributing to population declines of mountain hares in Scotland. We use hare data collected during red grouse counts in Scotland, to consider spatial and temporal variation in annual mountain hare indices of spring abundance in relation to different grouse management intensities, classified as either ‘Driven’ (driven grouse shooting), ‘Walked-up’ (walked-up shooting) or ‘Not-shot’ (where there was no grouse shooting interest). Trends in mountain hare abundance indices varied with region and grouse management intensity. Hare indices were higher and relatively stable on moors where driven grouse shooting was practised relative to lower indices and declines on moors where grouse were either walked-up or not shot. Abundance indices fluctuated over time in a quasi-cyclical manner, fluctuations being more pronounced where hares were more abundant, i.e. on driven grouse moors. Density-dependent processes such as resource competition and parasitism could explain greater fluctuations in areas where hare abundance indices were higher, whilst reductions in indices over time probably reflect sampling at decline phases of the cycle, particularly on driven moors, rather than being part of true long-term declines. Given our understanding of grouse moor management and mountain hare ecology, enhanced habitat management and control of generalist predators seem to be the most parsimonious explanation for higher abundances indices of mountain hare on driven grouse moors.

Current Status of the Eastern Sayan Snow Leopard (Panthera uncia) Grouping and Its Nutritive Base

Abstract—A field survey of snow leopard (Panthera uncia) habitats was carried out in the southeastern part of the Eastern Sayan Mountains (Okinskii and Tunkinskii districts of the Republic of Buryatia and the Kaa-Khemskii district of Tuva Republic). Seven or eight adult snow leopards were observed as constant inhabitants of the Tunkinskie Gol’tsy, Munku-Sardyk, and Bol’shoi Sayan mountain ridges. The presence of eight snow leopards was confirmed using DNA-based analyses of scats collected in 2014–2016. The main prey species of the snow leopard in Eastern Sayan is the Siberian ibex (Capra sibirica), but its abundance has steadily decreased over the past 20 years. The red deer (Cervus elaphus) and the wild boar (Sus scrofa), which were some of the most numerous ungulates in the survey area, are replacing the Siberian ibex in the snow leopard’s diet. In addition, the mountain hare (Lepus timidus) is also of importance to the snow leopard’s diet.

Overcoming species barriers: an outbreak of Lagovirus europaeus GI.2/RHDV2 in an isolated population of mountain hares (Lepus timidus)

BackgroundPrior to 2010, the lagoviruses that cause rabbit hemorrhagic disease (RHD) in European rabbits (Oryctolagus cuniculus) and European brown hare syndrome (EBHS) in hares (Lepus spp.) were generally genus-specific. However, in 2010, rabbit hemorrhagic disease virus 2 (RHDV2), also known as Lagovirus europaeus GI.2, emerged and had the distinguishing ability to cause disease in both rabbits and certain hare species. The mountain hare (Lepus timidus) is native to Sweden and is susceptible to European brown hare syndrome virus (EBHSV), also called Lagovirus europaeus GII.1. While most mountain hare populations are found on the mainland, isolated populations also exist on islands. Here we investigate a mortality event in mountain hares on the small island of Hallands Väderö where other leporid species, including rabbits, are absent.ResultsPost-mortem and microscopic examination of three mountain hare carcasses collected from early November 2016 to mid-March 2017 revealed acute hepatic necrosis consistent with pathogenic lagovirus infection. Using immunohistochemistry, lagoviral capsid antigen was visualized within lesions, both in hepatocytes and macrophages. Genotyping and immunotyping of the virus independently confirmed infection with L. europaeus GI.2, not GII.1. Phylogenetic analyses of the vp60 gene grouped mountain hare strains together with a rabbit strain from an outbreak of GI.2 in July 2016, collected approximately 50 km away on the mainland.ConclusionsThis is the first documented infection of GI.2 in mountain hares and further expands the host range of GI.2. Lesions and tissue distribution mimic those of GII.1 in mountain hares. The virus was most likely initially introduced from a concurrent, large-scale GI.2 outbreak in rabbits on the adjacent mainland, providing another example of how readily this virus can spread. The mortality event in mountain hares lasted for at least 4.5 months in the absence of rabbits, which would have required virus circulation among mountain hares, environmental persistence and/or multiple introductions. This marks the fourth Lepus species that can succumb to GI.2 infection, suggesting that susceptibility to GI.2 may be common in Lepus species. Measures to minimize the spread of GI.2 to vulnerable Lepus populations therefore are prudent.

Seasonal and predator-prey effects on circadian activity of free-ranging mammals revealed by camera traps.

Endogenous circadian and seasonal activity patterns are adapted to facilitate effective utilisation of environmental resources. Activity patterns are shaped by physiological constraints, evolutionary history, circadian and seasonal changes and may be influenced by other factors, including ecological competition and interspecific interactions. Remote-sensing camera traps allow the collection of species presence data throughout the 24 h period and for almost indefinite lengths of time. Here, we collate data from 10 separate camera trap surveys in order to describe circadian and seasonal activity patterns of 10 mammal species, and, in particular, to evaluate interspecific (dis)associations of five predator-prey pairs. We recorded 8,761 independent detections throughout Northern Ireland. Badgers, foxes, pine martens and wood mice were nocturnal; European and Irish hares and European rabbits were crepuscular; fallow deer and grey and red squirrels were diurnal. All species exhibited significant seasonal variation in activity relative to the timing of sunrise/sunset. Foxes in particular were more crepuscular from spring to autumn and hares more diurnal. Lagged regression analyses of predator-prey activity patterns between foxes and prey (hares, rabbits and wood mice), and pine marten and prey (squirrel and wood mice) revealed significant annual and seasonal cross-correlations. We found synchronised activity patterns between foxes and hares, rabbits and wood mice and pine marten and wood mice, and asynchrony between squirrels and pine martens. Here, we provide fundamental ecological data on endemic, invasive, pest and commercially valuable species in Ireland, as well as those of conservation importance and those that could harbour diseases of economic and/or zoonotic relevance. Our data will be valuable in informing the development of appropriate species-specific methodologies and processes and associated policies.

Post‐glacial range revolutions in South European hares (Lepus spp.): Insights from ancient DNA and ecological niche modelling

AbstractAimThe distribution of hares (Lepus spp.) in southern Europe was markedly different at the Last Glacial Maximum. Historical hybridization during range replacements led to high frequencies of mtDNA introgression from Lepus timidus into populations of three other species in northern Iberian Peninsula, even though L. timidus subsequently disappeared from the region. We use palaeontological records to provide new insights on the biogeographic history of these range replacements, combining ancient DNA and ecological niche modelling.LocationSouthern France.TaxonHares (genus Lepus).MethodsDNA was extracted from hare bones collected at Neolithic sites in southern France (5.5–7.5 thousand years before present; kyr bp) with uncertain species assignment. One mtDNA fragment was sequenced and 27 SNPs from 11 nuclear genes with species‐diagnostic information were genotyped. Distributions of L. granatensis, L. europaeus, and L. timidus were modelled using spatial and bioclimatic predictors, and the favourability function. Each model was transferred to 6 kyr bp and favourabilities were combined to determine the species with the highest environmental favourability in each sampled locality.ResultsAncient DNA analysis showed that the screened specimens belong to L. granatensis, a species presently confined to the Iberian Peninsula, but carried mtDNA haplotypes from L. timidus. Niche models show that L. granatensis achieved highest favourability in southern France 6 kyr bp reinforcing the molecular evidences.Main conclusionsL. granatensis replaced L. timidus populations from Iberia to southern France, being present in this region 5.5 kyr bp. Subsequent westwards invasion of L. europaeus likely replaced these L. granatensis populations towards northern Iberia, establishing current ranges. Despite successive species replacements, introgressed mtDNA haplotypes from L. timidus mark the distribution of the species in northern Iberian Peninsula before it disappeared in the region. Characterizing complex biogeographic histories of interacting species is a key to understand processes that led to current distributions of genetic diversity.

The importance of wildlife in the ecology and epidemiology of the TBE virus in Sweden: incidence of human TBE correlates with abundance of deer and hares

BackgroundTick-borne encephalitis (TBE) is one tick-transmitted disease where the human incidence has increased in some European regions during the last two decades. We aim to find the most important factors causing the increasing incidence of human TBE in Sweden. Based on a review of published data we presume that certain temperature-related variables and the population densities of transmission hosts, i.e. small mammals, and of primary tick maintenance hosts, i.e. cervids and lagomorphs, of the TBE virus vector Ixodes ricinus, are among the potentially most important factors affecting the TBE incidence. Therefore, we compare hunting data of the major tick maintenance hosts and two of their important predators, and four climatic variables with the annual numbers of human cases of neuroinvasive TBE. Data for six Swedish regions where human TBE incidence is high or has recently increased are examined by a time-series analysis. Results from the six regions are combined using a meta-analytical method.ResultsWith a one-year time lag, the roe deer (Capreolus capreolus), red deer (Cervus elaphus), mountain hare (Lepus timidus) and European hare (Lepus europaeus) showed positive covariance; the Eurasian elk (moose, Alces alces) and fallow deer (Dama dama) negative covariance; whereas the wild boar (Sus scrofa), lynx (Lynx lynx), red fox (Vulpes vulpes) and the four climate parameters showed no significant covariance with TBE incidence. All game species combined showed positive covariance.ConclusionsThe epidemiology of TBE varies with time and geography and depends on numerous factors, i.a. climate, virus genotypes, and densities of vectors, tick maintenance hosts and transmission hosts. This study suggests that the increased availability of deer to I. ricinus over large areas of potential tick habitats in southern Sweden increased the density and range of I. ricinus and created new TBEV foci, which resulted in increased incidence of human TBE. New foci may be established by TBE virus-infected birds, or by birds or migrating mammals infested with TBEV-infected ticks. Generally, persistence of TBE virus foci appears to require presence of transmission-competent small mammals, especially mice (Apodemus spp.) or bank voles (Myodes glareolus).

Seven decades of mountain hare counts show severe declines where high‐yield recreational game bird hunting is practised

Abstract Recreational hunting is widespread and can benefit nature conservation when well‐practised, monitored, and regulated. Management for recreational red grouse Lagopus lagopus scotica shooting on upland heathland in the UK causes conservation conflict because the intensive habitat, predator, and disease management needed to maintain high‐grouse densities for “driven” shooting has detrimental environmental impacts, notably for raptor populations. Sustainable management of mountain hares Lepus timidus scoticus, a game species in the same landscapes, poses a challenge. Control of transmission to grouse of a viral disease, louping‐ill, for which mountain hares are a host, has become an additional motivation to kill mountain hares since research during 1993–2001 suggested that culls might reduce infection rates in grouse. We analysed population trends of mountain hares from spring counts on moorland managed for grouse shooting and on contiguous alpine land. On moorland sites, a long‐term decline (4.6% per annum) from 1954 to 1999 increased to 30.7% per annum from then until 2017, with a density index falling to <1% of initial levels after 2008. Before 1999, declines were associated with conifer planting and were least severe where heather burning characteristic of grouse management was present. Grouse moors had the highest rate of decline after 1999. On alpine sites, the density index increased by 2.0% per annum from 1954 to 2007, then declined by 12.3% per annum but remained within the previous range of variation. Despite lack of evidence that it increases grouse numbers, reduction of louping‐ill transmission to grouse became a more frequent justification for mountain hare culls at a time consistent with it causing these recent, rapid mountain hare declines on grouse moors. Synthesis and applications. Long‐term field counts suggest that intensification of game bird management has resulted in severe, recent declines in mountain hare numbers, exacerbating longer term declines associated with land‐use change. Management practices founded on misinterpretation of earlier research are the probable cause. Regulation of hare culling would provide a framework for formal tests of whether culls affect grouse surpluses. It would also provide an opportunity to examine mountain hare populations' resilience to culls of varying size and seasonal timing.

Progressively excluding mammals of different body size affects community and trait structure of ground beetles

Mammalian grazing induces changes in vegetation properties in grasslands, which can affect a wide variety of other animals including many arthropods. However, the impacts may depend on the type and body size of these mammals. Furthermore, how mammals influence functional trait syndromes of arthropod communities is not well known.We progressively excluded large (e.g. red deer, chamois), medium (e.g. alpine marmot, mountain hare), and small (e.g. mice) mammals using size‐selective fences in two vegetation types (short‐ and tall‐grass vegetation) of subalpine grasslands. We then assessed how these exclusions affected the community composition and functional traits of ground beetles (Coleoptera, Carabidae), and which vegetation characteristic mediated the observed effects.Total carabid biomass, the activity densities of carabids with specific traits (i.e. small eyes, short wings), the richness of small‐eyed species and the richness of herbivorous species were significantly higher when certain mammals were excluded compared to when all mammals had access, regardless of vegetation type. Excluding large and medium mammals increased the activity density of herbivorous carabid species, but only in short‐grass vegetation. Similarly, excluding large mammals (ungulates) altered carabid species composition in the short‐, but not in the tall‐grass vegetation. All these responses were related to aboveground plant biomass, but not to plant Shannon diversity or vegetation structural heterogeneity.Our results indicate that changes in aboveground plant biomass are key drivers of mammalian grazers’ influence on carabids, suggesting that bottom–up forces are important in subalpine grassland systems. The exclusion of ungulates provoked the strongest carabid response. Our results, however, also highlight the ecological significance of smaller herbivorous mammals. Our study furthermore shows that mammalian grazing not only altered carabid community composition, but also caused community‐wide functional trait shifts, which could potentially have a wider impact on species interactions and ecosystem functioning.

Alpine glacial relict species losing out to climate change: The case of the fragmented mountain hare population (Lepus timidus) in the Alps.

Alpine and Arctic species are considered to be particularly vulnerable to climate change, which is expected to cause habitat loss, fragmentation and-ultimately-extinction of cold-adapted species. However, the impact of climate change on glacial relict populations is not well understood, and specific recommendations for adaptive conservation management are lacking. We focused on the mountain hare (Lepus timidus) as a model species and modelled species distribution in combination with patch and landscape-based connectivity metrics. They were derived from graph-theory models to quantify changes in species distribution and to estimate the current and future importance of habitat patches for overall population connectivity. Models were calibrated based on 1,046 locations of species presence distributed across three biogeographic regions in the Swiss Alps and extrapolated according to two IPCC scenarios of climate change (RCP 4.5 & 8.5), each represented by three downscaled global climate models. The models predicted an average habitat loss of 35% (22%-55%) by 2100, mainly due to an increase in temperature during the reproductive season. An increase in habitat fragmentation was reflected in a 43% decrease in patch size, a 17% increase in the number of habitat patches and a 34% increase in inter-patch distance. However, the predicted changes in habitat availability and connectivity varied considerably between biogeographic regions: Whereas the greatest habitat losses with an increase in inter-patch distance were predicted at the southern and northern edges of the species' Alpine distribution, the greatest increase in patch number and decrease in patch size is expected in the central Swiss Alps. Finally, both the number of isolated habitat patches and the number of patches crucial for maintaining the habitat network increased under the different variants of climate change. Focusing conservation action on the central Swiss Alps may help mitigate the predicted effects of climate change on population connectivity.

Mitochondrial DNA Introgression at the Northern Edge of the Brown Hare (Lepus europaeus) Range

Climate change enables expansion of southern species whereas many boreal species retreat northwards. Range shifts create novel interactions that can affect abundance and distribution via competition and/or hybridization. It is possible that the mountain hare (Lepus timidus) long-term decline in northern Europe is linked to competition with the brown hare (L. europaeus). In order to understand consequences of hare interactions at high latitudes, we determined geographical extent of hybridization using mitochondrial DNA (mtDNA) introgression as a proxy. Additionally, we compared species distribution and snow conditions in Finland with introgression frequency. We found that introgression is bidirectional but direction is mostly from the mountain hare to the brown hare. Approximately 10% of brown hares have introgressed mtDNA, being most common at the northern expansion front of their distribution range. Introgression is likely linked to brown hare expansion facilitated by climate change, but due to complex n...

Late Pleistocene hares of the Japanese archipelago: Paleobiogeographic implication at the Last Glacial Maximum

AbstractThe Tsugaru Strait is often referred to as Blakiston's Line in biogeography. Although this strait is thought to have become narrower during the Last Glacial Maximum, it did not close completely to form a land bridge. Sea‐level drawdown associated with cooling nevertheless weakened the influence of the Tsugaru Strait as a barrier to faunal migration. The currently accepted hypothesis is that only some large and highly mobile ungulates were able to migrate to Honshu over this strait to the exclusion of small mammals. However, this remains problematic as very few fossils corroborate this idea, and quantitative identification methods for use with tiny fragments of teeth have not been developed. In this study, we determine which of the species Lepus timidus (now distributed on Hokkaido and to the north) and Lepus brachyurus (now mainly distributed on Honshu, Shikoku, and Kyushu) can accurately be identified within a sample of Late Pleistocene Lepus sp. fossils recently unearthed from the Shitsukari‐Abe cave to the immediate south of the Tsugaru Strait. This is because we are unable to exclude the possibility that these fossils are referable to L. timidus on the basis of site location alone. Thus, utilising images of mandibular third premolar teeth obtained via microcomputed tomography scanning, we report (a) linear measurements, (b) a survey of the frequency of appearance of nonmetric traits, and (c) principal component analysis using landmark methods. The results show that fossil representatives of the genus Lepus can be differentiated from extant L. timidus and closely resemble L. brachyurus. Considering the past allopatric distribution of the 2 species suggested by other studies as well as fossil records around the Tsugaru Strait, this result corroborates the existing hypothesis that only large and highly mobile ungulates were able to navigate the Tsugaru Strait.

Widespread introgression of mountain hare genes into Fennoscandian brown hare populations.

In Fennoscandia, mountain hare (Lepus timidus) and brown hare (Lepus europaeus) hybridize and produce fertile offspring, resulting in gene flow across the species barrier. Analyses of maternally inherited mitochondrial DNA (mtDNA) show that introgression occur frequently, but unavailability of appropriate nuclear DNA markers has made it difficult to evaluate the scale- and significance for the species. The extent of introgression has become important as the brown hare is continuously expanding its range northward, at the apparent expense of the mountain hare, raising concerns about possible competition. We report here, based on analysis of 6833 SNP markers, that the introgression is highly asymmetrical in the direction of gene flow from mountain hare to brown hare, and that the levels of nuclear gene introgression are independent of mtDNA introgression. While it is possible that brown hares obtain locally adapted alleles from the resident mountain hares, the low levels of mountain hare alleles among allopatric brown hares suggest that hybridization is driven by stochastic processes. Interspecific geneflow with the brown hare is unlikely to have major impacts on mountain hare in Fennoscandia, but direct competition may.

Mountain hare transcriptome and diagnostic markers as resources to monitor hybridization with European hares

We report the first mountain hare (Lepus timidus) transcriptome, produced by de novo assembly of RNA-sequencing reads. Data were obtained from eight specimens sampled in two localities, Alps and Ireland. The mountain hare tends to be replaced by the invading European hare (Lepus europaeus) in their numerous contact zones where the species hybridize, which affects their gene pool to a yet unquantified degree. We characterize and annotate the mountain hare transcriptome, detect polymorphism in the two analysed populations and use previously published data on the European hare (three specimens, representing the European lineage of the species) to identify 4 672 putative diagnostic sites between the species. A subset of 85 random independent SNPs was successfully validated using PCR and Sanger sequencing. These valuable genomic resources can be used to design tools to assess population status and monitor hybridization between species.

The effects of acaricide treatment of sheep on red grouse Lagopus lagopus scotica tick burdens and productivity in a multi-host system.

Ixodes ricinus (Ixodida: Ixodidae) ticks are of economic and pathogenic importance across Europe. Within the uplands of the U.K., management to reduce ticks is undertaken to benefit red grouse Lagopus lagopus scotica (Galliformes: Phasianidae). Management strategies focus on the acaricide treatment of domestic sheep Ovis aries (Artiodactyla: Bovidae), but the effectiveness of this is less certain in the presence of wild hosts, particularly red deer Cervus elaphus (Artiodactyla: Cervidae) and mountain hare Lepus timidus (Lagomorpha: Leporidae). This study examines the effects of sheep management on grouse tick burdens and productivity using sites with a range of wild host densities. Sites at which applications of acaricide were more frequent had lower tick burdens; this relationship was similar on sites with a range of deer densities. However, no direct link was detected between acaricide treatment interval and grouse productivity. Sites with higher deer densities had higher grouse tick burdens and lower productivity [mean ± standard error (SE) young:adult ratio: 1.2 ± 0.2] compared with sites with lower deer densities (mean ± SE young:adult ratio: 1.8 ± 0.1). Sites with higher grouse brood sizes and higher proportions of hens with broods were also those with higher mountain hare abundance indices. This study highlights the importance of the frequent treatment of sheep with acaricide to reduce tick burdens on grouse, even in the presence of wild hosts.

積雪期におけるノウサギ・キツネの垂直分布と生息数 : 北海道石狩・留萌地方における

積雪期におけるノウサギ・キツネの垂直分布と生息数 : 北海道石狩・留萌地方における

Understanding hybridization and competition processes between hare species: Implications for conservation and management on the basis of a mathematical model

Hybridization and introgression frequently occur in animal communities, both in nature or caused by human actions. In the genus Lepus (Mammals, Lagomorphs) genetic mixing has been widely documented and it probably affected the population dynamics of species, particularly during climatic oscillations. We thus propose and investigate a mathematical model, with the aim to describe and predict the outcome of interspecific interactions in a three-population system including European (Lepus europaeus), mountain (Lepus timidus) and hybrid hares.Our results show that the three-population system can survive at a stable level, but equilibria involving only parental species could also be feasible. This implies that, depending on initial conditions, the mountain hare could suffer from the competition with European and hybrid hares and its population could disappear in the near future. Under climate change scenarios, the conditions for coexistence will become even more restrictive, mountain hare populations could be depressed and finally collapse.Thus, at present the hybridization phenomenon does not necessarily represent a threat to the conservation of the unique genomes of the parental species, but climate change and human intervention, namely the introduction of European hares in the range of the Alpine mountain species, could result in an increase in the contact zones between populations, finally promoting hybridization and biotic homogenization.

How genetic data improve the interpretation of results of faecal glucocorticoid metabolite measurements in a free-living population.

Measurement of glucocorticoid metabolites (GCM) in faeces has become a widely used and effective tool for evaluating the amount of stress experienced by animals. However, the potential sampling bias resulting from an oversampling of individuals when collecting “anonymous” (unknown sex or individual) faeces has rarely been investigated. We used non-invasive genetic sampling (NIGS) to investigate potential interpretation errors of GCM measurements in a free-living population of mountain hares during the mating and post-reproductive periods. Genetic data improved the interpretation of results of faecal GCM measurements. In general GCM concentrations were influenced by season. However, genetic information revealed that it was sex-dependent. Within the mating period, females had higher GCM levels than males, but individual differences were more expressed in males. In the post-reproductive period, GCM concentrations were neither influenced by sex nor individual. We also identified potential pitfalls in the interpretation of anonymous faecal samples by individual differences in GCM concentrations and resampling rates. Our study showed that sex- and individual-dependent GCM levels led to a misinterpretation of GCM values when collecting “anonymous” faeces. To accurately evaluate the amount of stress experienced by free-living animals using faecal GCM measurements, we recommend documenting individuals and their sex of the sampled population. In stress-sensitive and elusive species, such documentation can be achieved by using NIGS and for diurnal animals with sexual and individual variation in appearance or marked individuals, it can be provided by a detailed field protocol.

Tularaemia in European Brown Hares (Lepus europaeus) and Mountain Hares (Lepus timidus) Characterized by Histopathology and Immunohistochemistry: Organ Lesions and Suggestions of Routes of Infection and Shedding

Tularaemia is an emerging zoonotic infectious disease caused by the bacterium Francisella tularensis. In Sweden, hares are considered to be key species in the epidemiology of tularaemia. The aim of this study was to characterize the pathology of natural tularaemia infection in European brown hares (EBHs) (Lepus europaeus) and mountain hares (MHs) (Lepus timidus) in Sweden, in order to better understand the presentation of disease and the routes of infection, body dissemination and shedding of F. tularensis. During 2000-2013, 49 EBHs and 37 MHs were diagnosed with tularaemia. Enlargement of the spleen was seen in 80% of EBHs and 62% of MHs. Necrosis was often obvious in the bone marrow, liver, lung and spleen, but 30% of the hares had no lesions or minimal gross lesions. On microscopical examination of tissues from 27 EBHs and three MHs, necrosis was seen in the majority of samples of liver, spleen, bone marrow, lymph node and adrenal glands and was common in the lungs and brain meninges. Immunohistochemistry for Francisella spp. detected bacteria in association with necrosis and inflammation. In several cases, Francisella spp. were also found inside blood vessels, in the renal pelvis, in lactating mammary glands, in bronchioles and in the skin, associated with tick bites. Using quantitative polymerase chain reaction, two genotypes of F. tularensis subsp. holarctica were found; canSNP group B.6, all belonging to subgroup B.7, and canSNP group B.12. There were no differences in pathology between the genotypes. Our results indicate that the urinary tract and mammary glands are important routes for the shedding of F. tularensis. Hunters may not be aware of the risks of contracting tularaemia while handling hares, since infected hares do not always show noticeable gross lesions.

A single multiplex of twelve microsatellite markers for the simultaneous study of the brown hare (Lepus europaeus) and the mountain hare (Lepus timidus)

AbstractThe management of hunted species is challenging, as it must conciliate the conservation of species and their sustainable exploitation. Nongenetic tools are widely used in this context but they may present limitations notably when species can hybridize or when large‐scale spatial monitoring is required to establish optimal management actions. This is why genetic tools have been more and more integrated in wildlife management practices. However, the markers proposed are often amplified in small multiplexes when larger ones could allow to better cope with the small quantities of DNA obtained with noninvasive sampling methods. Here, we propose a unique multiplex of 12 autosomal microsatellite markers for the study of two hare species that exist in sympatry in some areas in Europe and are hunted notably in France: the brown hare Lepus europaeus and the mountain hare L. timidus. We tested 17 markers previously used in these two species or other lagomorph species, from which 12 were included in this single multiplex. Diversity was between 4 and 30 alleles per locus totaling 126 alleles, and we showed that these markers possess appropriate genetic resolution for individual and species identification for the populations under study. This multiplex panel represents the largest number of microsatellites amplified in one reaction proposed for these two hare species and provides a cost‐effective and valuable tool for further hybridization studies and the management of hares.