From Caves to the Savannah, the Mitogenome History of Modern Lions (Panthera leo) and Their Ancestors.

Lions (Panthera leo) play a crucial ecological role in shaping and maintaining fragile ecosystems within Africa. Conservation efforts should focus on genetic variability within wild populations when considering reintroduction attempts. We studied two groups of lions from two conservation sites located in Zambia and Zimbabwe to determine their genetic make-up, information that is usually unknown to the sites. In this study, we analysed 17 specimens for cytb and seven microsatellite markers to ascertain family relationships and genetic diversity previously obtained by observational studies. We then produced a standardised haplogroup phylogeny using all available entire mitogenomes, as well as calculating a revised molecular clock. The modern lion lineage diverged ~151 kya and was divided into two subspecies, both containing three distinct haplogroups. We confirm that Panthera leo persica is not a subspecies, but rather a haplogroup of the northern P.l. leo that exited Africa at least ~31 kya. The progenitor to all lions existed ~1.2 Mya, possibly in SE Africa, and later exited Africa and split into the two cave lion lineages ~175 kya. Species demography is correlated to major climactic events. We now have a detailed phylogeny of lion evolution and an idea of their conservation status given the threat of climate change.

African or Asiatic origin? Genome analyses solved the mystery of the hybrid origin of the rescued lion cub.

Species assignment of any seized material using DNA analysis has been a routine and widely accepted standard procedure in providing scientific advisory for the legal prosecution of wildlife cases. Scientific advancements and rigorous application of genetic tools have led to the development of a variety of molecular markers with their defined efficacy in wildlife forensics. However, in a few unusual cases where a hybrid needs to be identified or assignment need to be made at sub-species level, mitochondrial markers often fail or else provide biased results, which can affect the overall judgment in the court of law. Here, we report one such challenging case of lion cub rescued by the law enforcement from illegal trafficking. Phylogenetic assessment based on complete mitogenome assigned rescued lion cub with African lion (Panthera leo leo). However, the TSPY gene of the Y chromosome established that the lion cub shared its paternal lineage from Asiatic lion (Panthera leo persica). With the use of maternally and paternally inherited markers, we conclude a hybrid origin of the rescued lion cub which shared ancestry from both Asiatic as well as African lion. The present study exhibits the application of genome sequencing in thinking beyond routine identification and contributes to the operating procedures of wildlife forensics.

The evolutionary history of extinct and living lions.

Lions are one of the world's most iconic megafauna, yet little is known about their temporal and spatial demographic history and population differentiation. We analyzed a genomic dataset of 20 specimens: two ca. 30,000-y-old cave lions (Panthera leo spelaea), 12 historic lions (Panthera leo leo/Panthera leo melanochaita) that lived between the 15th and 20th centuries outside the current geographic distribution of lions, and 6 present-day lions from Africa and India. We found that cave and modern lions shared an ancestor ca. 500,000 y ago and that the 2 lineages likely did not hybridize following their divergence. Within modern lions, we found 2 main lineages that diverged ca. 70,000 y ago, with clear evidence of subsequent gene flow. Our data also reveal a nearly complete absence of genetic diversity within Indian lions, probably due to well-documented extremely low effective population sizes in the recent past. Our results contribute toward the understanding of the evolutionary history of lions and complement conservation efforts to protect the diversity of this vulnerable species.

Long live the king: chromosome-level assembly of the lion (Panthera leo) using linked-read, Hi-C, and long-read data.

BACKGROUND: The lion (Panthera leo) is one of the most popular and iconic feline species on the planet, yet in spite of its popularity, the last century has seen massive declines for lion populations worldwide. Genomic resources for endangered species represent an important way forward for the field of conservation, enabling high-resolution studies of demography, disease, and population dynamics. Here, we present a chromosome-level assembly from a captive African lion from the Exotic Feline Rescue Center (Center Point, IN) as a resource for current and subsequent genetic work of the sole social species of the Panthera clade. RESULTS: Our assembly is composed of 10x Genomics Chromium data, Dovetail Hi-C, and Oxford Nanopore long-read data. Synteny is highly conserved between the lion, other Panthera genomes, and the domestic cat. We find variability in the length of runs of homozygosity across lion genomes, indicating contrasting histories of recent and possibly intense inbreeding and bottleneck events. Demographic analyses reveal similar ancient histories across all individuals during the Pleistocene except the Asiatic lion, which shows a more rapid decline in population size. We show a substantial influence on the reference genome choice in the inference of demographic history and heterozygosity. CONCLUSIONS: We demonstrate that the choice of reference genome is important when comparing heterozygosity estimates across species and those inferred from different references should not be compared to each other. In addition, estimates of heterozygosity or the amount or length of runs of homozygosity should not be taken as reflective of a species, as these can differ substantially among individuals. This high-quality genome will greatly aid in the continuing research and conservation efforts for the lion, which is rapidly moving towards becoming a species in danger of extinction.

A genome-wide data assessment of the African lion (Panthera leo) population genetic structure and diversity in Tanzania.

The African lion (Panthera leo), listed as a vulnerable species on the IUCN Red List of Threatened Species (Appendix II of CITES), is mainly impacted by indiscriminate killing and prey base depletion. Additionally, habitat loss by land degradation and conversion has led to the isolation of some subpopulations, potentially decreasing gene flow and increasing inbreeding depression risks. Genetic drift resulting from weakened connectivity between strongholds can affect the genetic health of the species. In the present study, we investigated the evolutionary history of the species at different spatiotemporal scales. Therefore, the mitochondrial cytochrome b gene (N = 128), 11 microsatellites (N = 103) and 9,103 SNPs (N = 66) were investigated in the present study, including a large sampling from Tanzania, which hosts the largest lion population among all African lion range countries. Our results add support that the species is structured into two lineages at the continental scale (West-Central vs East-Southern), underlining the importance of reviewing the taxonomic status of the African lion. Moreover, SNPs led to the identification of three lion clusters in Tanzania, whose geographical distributions are in the northern, southern and western regions. Furthermore, Tanzanian lion populations were shown to display good levels of genetic diversity with limited signs of inbreeding. However, their population sizes seem to have gradually decreased in recent decades. The highlighted Tanzanian African lion population genetic differentiation appears to have resulted from the combined effects of anthropogenic pressure and environmental/climatic factors, as further discussed.

Dietary behaviour of man-eating lions as revealed by dental microwear textures.

Lions (Panthera leo) feed on diverse prey species, a range that is broadened by their cooperative hunting. Although humans are not typical prey, habitual man-eating by lions is well documented. Fathoming the motivations of the Tsavo and Mfuwe man-eaters (killed in 1898 in Kenya and 1991 in Zambia, respectively) may be elusive, but we can clarify aspects of their behaviour using dental microwear texture analysis. Specifically, we analysed the surface textures of lion teeth to assess whether these notorious man-eating lions scavenged carcasses during their depredations. Compared to wild-caught lions elsewhere in Africa and other large feliforms, including cheetahs and hyenas, dental microwear textures of the man-eaters do not suggest extreme durophagy (e.g. bone processing) shortly before death. Dental injuries to two of the three man-eaters examined may have induced shifts in feeding onto softer foods. Further, prompt carcass reclamation by humans likely limited the man-eaters' access to bones. Man-eating was likely a viable alternative to hunting and/or scavenging ungulates due to dental disease and/or limited prey availability.

Retrospective study of mortality in Asiatic lions (Panthera leo persica) in the European breeding population between 2000 and 2014.

Although the European population of Asiatic lions (Panthera leo persica) has been managed under the European Endangered Species Program (EEP) since 1990, little is known about the health status of the population. This study was designed to characterize mortality for this population through examination of the studbook and other records on 392 Asiatic lions living in the EEP between 2000 and 2014. A total of 270 animals died during the period with 80% of them being under 1 year old. The mortality rate for under 1 year olds was 54%, while the odds of survival of cubs within a litter increased if the dam had had litters previously. Survival to reproductive age was 44%. Post-mortem reports were requested and the cause of death was obtained for 133 animals. Trauma inflicted by a conspecific and lack of care were common causes of death (26% and 22%, respectively) and were also responsible for most of the neonatal mortalities. Congenital defects were responsible for 9% of deaths, although the true prevalence is likely underestimated. A common necropsy protocol for all Asiatic lion collections is needed to facilitate future studies. Zoo Biol. 36:66-73, 2017. © 2017 Wiley Periodicals, Inc.

Phylogeographic Patterns in Africa and High Resolution Delineation of Genetic Clades in the Lion (Panthera leo).

Comparative phylogeography of African savannah mammals shows a congruent pattern in which populations in West/Central Africa are distinct from populations in East/Southern Africa. However, for the lion, all African populations are currently classified as a single subspecies (Panthera leo leo), while the only remaining population in Asia is considered to be distinct (Panthera leo persica). This distinction is disputed both by morphological and genetic data. In this study we introduce the lion as a model for African phylogeography. Analyses of mtDNA sequences reveal six supported clades and a strongly supported ancestral dichotomy with northern populations (West Africa, Central Africa, North Africa/Asia) on one branch, and southern populations (North East Africa, East/Southern Africa and South West Africa) on the other. We review taxonomies and phylogenies of other large savannah mammals, illustrating that similar clades are found in other species. The described phylogeographic pattern is considered in relation to large scale environmental changes in Africa over the past 300,000 years, attributable to climate. Refugial areas, predicted by climate envelope models, further confirm the observed pattern. We support the revision of current lion taxonomy, as recognition of a northern and a southern subspecies is more parsimonious with the evolutionary history of the lion.

Mitochondrial Haplotype Diversity in Zambian Lions: Bridging a Gap in the Biogeography of an Iconic Species.

Analysis of DNA sequence diversity at the 12S to 16S mitochondrial genes of 165 African lions (Panthera leo) from five main areas in Zambia has uncovered haplotypes which link Southern Africa with East Africa. Phylogenetic analysis suggests Zambia may serve as a bridge connecting the lion populations in southern Africa to eastern Africa, supporting earlier hypotheses that eastern-southern Africa may represent the evolutionary cradle for the species. Overall gene diversity throughout the Zambian lion population was 0.7319 +/- 0.0174 with eight haplotypes found; three haplotypes previously described and the remaining five novel. The addition of these five novel haplotypes, so far only found within Zambia, nearly doubles the number of haplotypes previously reported for any given geographic location of wild lions. However, based on an AMOVA analysis of these haplotypes, there is little to no matrilineal gene flow (Fst = 0.47) when the eastern and western regions of Zambia are considered as two regional sub-populations. Crossover haplotypes (H9, H11, and Z1) appear in both populations as rare in one but common in the other. This pattern is a possible result of the lion mating system in which predominately males disperse, as all individuals with crossover haplotypes were male. The determination and characterization of lion sub-populations, such as done in this study for Zambia, represent a higher-resolution of knowledge regarding both the genetic health and connectivity of lion populations, which can serve to inform conservation and management of this iconic species.

Revealing the maternal demographic history of Panthera leo using ancient DNA and a spatially explicit genealogical analysis.

BACKGROUND: Understanding the demographic history of a population is critical to conservation and to our broader understanding of evolutionary processes. For many tropical large mammals, however, this aim is confounded by the absence of fossil material and by the misleading signal obtained from genetic data of recently fragmented and isolated populations. This is particularly true for the lion which as a consequence of millennia of human persecution, has large gaps in its natural distribution and several recently extinct populations. RESULTS: We sequenced mitochondrial DNA from museum-preserved individuals, including the extinct Barbary lion (Panthera leo leo) and Iranian lion (P. l. persica), as well as lions from West and Central Africa. We added these to a broader sample of lion sequences, resulting in a data set spanning the historical range of lions. Our Bayesian phylogeographical analyses provide evidence for highly supported, reciprocally monophyletic lion clades. Using a molecular clock, we estimated that recent lion lineages began to diverge in the Late Pleistocene. Expanding equatorial rainforest probably separated lions in South and East Africa from other populations. West African lions then expanded into Central Africa during periods of rainforest contraction. Lastly, we found evidence of two separate incursions into Asia from North Africa, first into India and later into the Middle East. CONCLUSIONS: We have identified deep, well-supported splits within the mitochondrial phylogeny of African lions, arguing for recognition of some regional populations as worthy of independent conservation. More morphological and nuclear DNA data are now needed to test these subdivisions.

First Asian record of Panthera (Leo) fossilis (Mammalia, Carnivora, Felidae) in the Early Pleistocene of Western Siberia, Russia.

A lion-like pantherine felid is described as Panthera (Leo) fossilis from the late Early Pleistocene sediments of the Kuznetsk Basin (Western Siberia, Russia). The find of P. fossilis first recorded in Asia considerably extends the current notion of the eastward expansion of the most ancient lions. The Siberian lion is geologically the oldest form and is dimensionally among the largest members of the group of fossil lions on the Eurasian continent. Although known by mandibular remains only, it is readily distinguished from Panthera (Leo) spelaea by a heavy built mandibular corpus with rectangular profile in the cheek teeth area, a deep, well-outlined and narrow anterior section of the masseteric fossa, and a large p4 supported by a big unreduced anterior root. The Siberian lion shares these features with the European Middle Pleistocene P. fossilis and the American Late Pleistocene P. (Leo) atrox, which suggests their close relationship. P. atrox originated from P. fossilis and was isolated in North America south of the Late Pleistocene ice sheets. This explains why the American lion has retained more primitive features than the coeval Eurasian cave lion P. (L.) spelaea.

The clavicles of Smilodon fatalis and Panthera atrox (Mammalia: Felidae) from Rancho La Brea, Los Angeles, California.

The Rancho La Brea collections at the George C. Page Museum in Los Angeles, California, contain the largest single inventory of Smilodon fatalis remains representing virtually every bone in the skeleton. Eighteen clavicles of two distinctive shapes have been recovered from historical and recent excavations at Rancho La Brea. In this study, we identify these specimens to species through comparison of their morphology and morphological variability with clavicles found in modern felids. This study includes a reevaluation of clavicles that have been previously assigned to S. fatalis, which are more likely to be those of Panthera atrox, and the description of pantherine cat clavicles. A previously undescribed sample of clavicles not only includes some of the same pantherine morph but also 10 specimens, herein assigned to S. fatalis, which are morphologically distinctive and significantly smaller than the previously described specimens. In addition, we report unexpected variations between clavicles of Panthera leo and P. tigris: the clavicles of P. leo closely resemble those of the large Rancho La Brea clavicle morph-which presumably belongs to P. atrox-thus supporting a P. leo/P. atrox clade. We report distinctive morphology of the clavicles of Acinonyx jubatus. Possible functional and phylogenic significance of felid clavicles is suggested.

Contacts between domestic livestock and wildlife at the Kruger National Park Interface of the Republic of South Africa.

One of the most important transboundary animal diseases (TADs) in the southern African region is foot-and-mouth disease (FMD). In this region, a pathway for spread of FMD virus is contacts between cattle and certain species of wildlife. The objective of this study was to evaluate contacts between cattle and wildlife in the Kruger National Park (KNP) and the adjacent Limpopo province for the time periods October 2006 to March 2007 and April to September 2007. In this study, 87 livestock owners and 57 KNP field rangers were interviewed. Fifteen (17%) livestock owners reported contacts between wildlife and cattle. More livestock owners reported observing contacts between cattle and all wildlife species during October-March than April-September (p=0.012). However, no difference was found between these periods for contacts between cattle and individual wildlife species. A total of 18 (32%) field rangers reported contacts between cattle and wildlife. The most common species-specific contacts were between cattle and buffalo (63/year), cattle and impala (17/year) and cattle and lion (10/year). There were no significant differences in rangers reporting observed contacts between cattle and wildlife during October-March versus April-September or between rangers reporting observed contacts outside versus within the KNP. Overall, there was no evidence of higher contact rates between cattle and wildlife in the study area during October-March compared to April-September. Contact data collected in this study can be used to better understand the transmission of FMD virus in this region.

Phylogeography of lions (Panthera leo ssp.) reveals three distinct taxa and a late Pleistocene reduction in genetic diversity.

Lions were the most widespread carnivores in the late Pleistocene, ranging from southern Africa to the southern USA, but little is known about the evolutionary relationships among these Pleistocene populations or the dynamics that led to their extinction. Using ancient DNA techniques, we obtained mitochondrial sequences from 52 individuals sampled across the present and former range of lions. Phylogenetic analysis revealed three distinct clusters: (i) modern lions, Panthera leo; (ii) extinct Pleistocene cave lions, which formed a homogeneous population extending from Europe across Beringia (Siberia, Alaska and western Canada); and (iii) extinct American lions, which formed a separate population south of the Pleistocene ice sheets. The American lion appears to have become genetically isolated around 340 000 years ago, despite the apparent lack of significant barriers to gene flow with Beringian populations through much of the late Pleistocene. We found potential evidence of a severe population bottleneck in the cave lion during the previous interstadial, sometime after 48 000 years, adding to evidence from bison, mammoths, horses and brown bears that megafaunal populations underwent major genetic alterations throughout the last interstadial, potentially presaging the processes involved in the subsequent end-Pleistocene mass extinctions.

The evolutionary dynamics of the lion Panthera leo revealed by host and viral population genomics.

The lion Panthera leo is one of the world's most charismatic carnivores and is one of Africa's key predators. Here, we used a large dataset from 357 lions comprehending 1.13 megabases of sequence data and genotypes from 22 microsatellite loci to characterize its recent evolutionary history. Patterns of molecular genetic variation in multiple maternal (mtDNA), paternal (Y-chromosome), and biparental nuclear (nDNA) genetic markers were compared with patterns of sequence and subtype variation of the lion feline immunodeficiency virus (FIV(Ple)), a lentivirus analogous to human immunodeficiency virus (HIV). In spite of the ability of lions to disperse long distances, patterns of lion genetic diversity suggest substantial population subdivision (mtDNA Phi(ST) = 0.92; nDNA F(ST) = 0.18), and reduced gene flow, which, along with large differences in sero-prevalence of six distinct FIV(Ple) subtypes among lion populations, refute the hypothesis that African lions consist of a single panmictic population. Our results suggest that extant lion populations derive from several Pleistocene refugia in East and Southern Africa ( approximately 324,000-169,000 years ago), which expanded during the Late Pleistocene ( approximately 100,000 years ago) into Central and North Africa and into Asia. During the Pleistocene/Holocene transition ( approximately 14,000-7,000 years), another expansion occurred from southern refugia northwards towards East Africa, causing population interbreeding. In particular, lion and FIV(Ple) variation affirms that the large, well-studied lion population occupying the greater Serengeti Ecosystem is derived from three distinct populations that admixed recently.

The origin, current diversity and future conservation of the modern lion (Panthera leo).

Understanding the phylogeographic processes affecting endangered species is crucial both to interpreting their evolutionary history and to the establishment of conservation strategies. Lions provide a key opportunity to explore such processes; however, a lack of genetic diversity and shortage of suitable samples has until now hindered such investigation. We used mitochondrial control region DNA (mtDNA) sequences to investigate the phylogeographic history of modern lions, using samples from across their entire range. We find the sub-Saharan African lions are basal among modern lions, supporting a single African origin model of modern lion evolution, equivalent to the 'recent African origin' model of modern human evolution. We also find the greatest variety of mtDNA haplotypes in the centre of Africa, which may be due to the distribution of physical barriers and continental-scale habitat changes caused by Pleistocene glacial oscillations. Our results suggest that the modern lion may currently consist of three geographic populations on the basis of their recent evolutionary history: North African-Asian, southern African and middle African. Future conservation strategies should take these evolutionary subdivisions into consideration.