Patterns of molecular genetic variation among African elephant populations

The highly threatened African elephants have recently been subdivided into two species, Loxodonta africana (savannah or bush elephant) and L. cyclotis (forest elephant) based on morphological and molecular studies. A molecular genetic assessment of 16 microsatellite loci across 20 populations (189 individuals) affirms species level genetic differentiation and provides robust genotypic assessment of species affiliation. Savannah elephant populations show modest levels of phylogeographic subdivision based on composite microsatellite genotype, an indication of recent population isolation and restricted gene flow between locales. The savannah elephants show significantly lower genetic diversity than forest elephants, probably reflecting a founder effect in the recent history of the savannah species.     

The evolution and phylogeography of the African elephant inferred from mitochondrial DNA sequence and nuclear microsatellite markers

Recent genetic results support the recognition of two African elephant species: Loxodonta africana, the savannah elephant, and Loxodonta cyclotis, the forest elephant. The study, however, did not include the populations of West Africa, where the taxonomic affinities of elephants have been much debated. We examined mitochondrial cytochrome b control region sequences and four microsatellite loci to investigate the genetic differences between the forest and savannah elephants of West and Central Africa. We then combined our data with published control region sequences from across Africa to examine patterns at the continental level. Our analysis reveals several deeply divergent lineages that do not correspond with the currently recognized taxonomy: (i) the forest elephants of Central Africa; the forest and savannah elephants of West Africa; and (iii) the savannah elephants of eastern, southern and Central Africa. We propose that the complex phylogeographic patterns we detect in African elephants result from repeated continental-scale climatic changes over their five-to-six million year evolutionary history. Until there is consensus on the taxonomy, we suggest that the genetic and ecological distinctness of these lineages should be an important factor in conservation management planning.     

Contribution of molecular phylogeny and morphometrics to the systematics of African elephants

African elephants are conventionally classified as a single species: Loxodonta africana (Blumenbach 1797). However, the discovery in 1900 of a smaller form of the African elephant, spread throughout the equatorial belt of this land, has given rise to a debate over the relevance of a second species of elephant in Africa. The twentieth century has not provided any definite answer to this question. Actually, recent molecular analyses have sustained this issue by advocating either a division of forest elephants into a valid species, or their inclusion as a subspecies of L. africana. Our work initiated at the National Museum of Natural History of Paris provides new molecular (mitochondrial) and morphological (and morphometrical) evidence making it possible to propose a comprehensive phylogenetic hypothesis. It appears that there is no conclusive argument to keep forest elephants (cyclotis form) and savannah elephants (africana form) apart in two distinct species. A high level of mitochondrial introgression between the two forms, as well as a continuum in the morphology of the skulls of the two morphotypes rather suggests that, despite an ancient division, these two taxa freely interbreed wherever their ranges intersect. We thus adopt a conservative systematic position in considering these two forms as two subspecies, respectively: L. africana africana, the savannah elephant, and L. africana cyclotis, the forest elephant. We finally discuss the conservation topic in the light of this systematic framework.     

Relatedness and demography of African forest elephants: inferences from noninvasive fecal DNA analyses

African forest elephants (Loxodonta cyclotis) are genetically and morphologically distinct from their savannah counterparts, but their biology remains poorly understood. In this study, I use noninvasive fecal DNA analyses to examine the relatedness structure and historical demography of forest elephants at 2 sites in SW Gabon, central Africa. Pairwise relatedness values calculated between 162 elephant individuals genotyped at 8 microsatellite loci were significantly higher within spatially associated dung piles than between random pairings for one site. First- and second-order relatives were most commonly detected among dung piles from adult female pairs and adult females and juveniles. Pairwise relatedness estimates suggested that, like savannah elephants, forest groups are largely composed of adult females, their sisters, and juvenile offspring. Associations between males, and groups containing juveniles from multiple related females, were detected but at much lower frequency. Analysis of mitochondrial d-loop sequences from 70 elephant individuals identified 2 haplogroups in SW Gabon.     

Characterization of tetranucleotide microsatellite loci in the African Savannah Elephant (Loxodonta africana africana)

Most African elephant (Loxodonta africana africana) populations are isolated and thus threatened by a loss of genetic diversity. As a consequence, genetic analysis of African elephant populations will play an increasing role in their conservation, and microsatellite loci will be an important tool in these analyses. Previously published sets of polymorphic microsatellites developed for African elephants are all dinucleotide repeats, which are prone to typing error. Here, we characterize 11 tetranucleotide microsatellite loci in the African elephant. All loci were polymorphic in 32 faecal samples and two tissue samples from 33 individual African savannah elephants.     

Sacoglottis gabonensis– a keystone fruit for forest elephants in the Réserve de Faune du Petit Loango, Gabon

This study suggests that the fruits of Sacoglottis gabonensis (Baill.) Urb. (Humiriaceae) are a keystone resource for forest elephants ( Loxodonta cyclotis Matschie) in a coastal rain forest, the Réserve de Faune du Petit Loango, Gabon (now part of Loango National Park). Faecal counts demonstrated that forest elephants used Sacoglottis -dominated forest more when Sacoglottis was abundant and electivity indices suggest that Sacoglottis is a preferred food. The flora of Petit Loango is characterized by the absence of herbaceous vegetation such as Marantaceae and Zingiberaceae, and during the prolonged dry season few fleshy fruits are present other than Sacoglottis fruits, which are produced in a glut during this time. While inter-annual fruiting reliability remains to be confirmed, fruit production in 1998 and high stem density relative to other study sites provide indirect evidence that Sacoglottis fruits are a reliable inter-annual resource at Petit Loango. It is thus proposed that Sacoglottis gabonensis fruits fulfil an important role as a keystone 'fallback' resource for forest elephants during the dry season at Petit Loango.     

Molecular phylogeny of Elephantidae. Extreme divergence of the extant forest African elephant.

A phylogenetic study of the Elephantidae (Proboscidea, Mammalia) is based on the cytochrome b mitochondrial gene: 31 terminals, that is, all known sequences, one non-elephantid proboscidean, the extinct American mastodon, and four outgroups. The data set includes 11 new sequences with the first published sequence of the forest African elephant, L. a. cyclotis. The analyses of extant taxa only and of both extant and extinct taxa show that L. a. cyclotis is highly divergent from L. a. africana. It is as divergent from L. a. africana as Loxodonta is divergent from Elephas. Southern L. a. africana form a clade. The continental subspecies E. m. indicus is paraphyletic with individuals from India and Thailand closer to E. m. maximus (Sri-Lanka). Members of Mammuthus primigenius are more closely related to Loxodonta although they do not form a clade; two specimens of M. primigenius are closer to L. a. africana making the genus Loxodonta paraphyletic. The latter conclusion may be partly due to unequal length of the various polymorphic mammoth sequences.     

Genetic identification of five strongyle nematode parasites in wild african elephants (Loxodonta africana)

African savannah elephants (Loxodonta africana) are an ecologically and economically important species in many African habitats. However, despite the importance of elephants, research on their parasites is limited, especially in wild populations. Currently, we lack genetic tools to identify elephant parasites. We present genetic markers from ribosomal DNA (rDNA) and mitochondrial DNA (mtDNA) to identify five elephant-specific nematode parasites in the family Strongylidae: Murshidia linstowi, Murshidia longicaudata, Murshidia africana, Quilonia africana, and Khalilia sameera. We collected adult nematodes from feces deposited by wild elephants living in Amboseli National Park, Kenya. Using both morphologic and genetic techniques, we found that the internal transcribed spacer (ITS) region in rDNA provides a reliable marker to distinguish these species of strongyles. We found no evidence for cryptic genetic species within these morphologic species according to the cox-1 region of mtDNA. Levels of genetic diversity in strongyles from elephants were consistent with the genetic diversity seen within other strongyle species. We anticipate that these results will be a useful tool for identifying gastrointestinal nematode parasites in elephants.     

Identifying Source Populations and Genetic Structure for Savannah Elephants in Human-Dominated Landscapes and Protected Areas in the Kenya-Tanzania Borderlands

We investigated the genetic metapopulation structure of elephants across the trans Rift Valley region of Kenya and Tanzania, one of the remaining strongholds for savannah elephants (Loxodonata africana) in East Africa, using microsatellite and mitochondrial DNA (mtDNA) markers. We then examined this population structure to determine the source population for a recent colonization event of savannah elephants on community-owned land within the trans rift valley region. Four of the five sampled populations showed significant genetic differentiation (p<0.05) as measured with both mtDNA haplotypes and microsatellites. Only the samples from the adjacent Maasai Mara and Serengeti ecosystems showed no significant differentiation. A phylogenetic neighbour-joining tree constructed from mtDNA haplotypes detected four clades. Clade four corresponds to the F clade of previous mtDNA studies that reported to have originated in forest elephants (Loxodonta cyclotis) but to also be present in some savannah elephant populations. The split between clade four and the other three clades corresponded strongly to the geographic distribution of mtDNA haplotypes across the rift valley in the study area. Clade four was the dominant clade detected on the west side of the rift valley with rare occurrences on the east side. Finally, the strong patterns of population differentiation clearly indicated that the recent colonists to the community-owned land in Kenya came from the west side of the rift valley. Our results indicate strong female philopatry within the isolated populations of the trans rift valley region, with gene flow primarily mediated via male movements. The recent colonization event from Maasai Mara or Serengeti suggests there is hope for maintaining connectivity and population viability outside formal protected areas in the region.     

A case study of apparent conflict between molecular phylogenies: the interrelationships of African elephants

Recent molecular phylogenies of the African elephants suggest that there is an evolutionary structure within Loxodonta africana. Some nuclear results ( Roca et al., 2001 ) support the separation of the forest African elephant subspecies L. a. cyclotis as a species distinct from the savannah elephant L. a. africana, on the basis of the recognition of both forming highly divergent (reciprocally monophyletic) clades. Conversely, a mitochondrial survey ( Eggert et al., 2002 ), while admitting a geographic partitioning of the genetic structure within African elephants, suggests retaining the status quo. They recognize three diagnosible entities (western, central and south‐eastern Africa) with non‐overlapping ranges within L. africana sensu lato. In order to address these conflicting views (historical fragmentation and speciation or isolation by distance, respectively), we have sequenced two datasets of 1961 bp (for 50 elephants) and about 3700 bp, respectively (for 20 elephants) of the mitochondrial DNA for both forms of elephants (cyclotis and africana). They span the cytochrome b gene, the control region and several RNAs. When compared with former mtDNA data, they provide the most comprehensive view of the African elephant phylogeny (78 mtDNA haplotypes, of which 44 are new) and provide the first insight into populations from the Democratic Republic of Congo. The genetic diversity of mtDNA was appraised and the stability of alternative phylogenetic trees was investigated. Our results are inconsistent with both those prior studies. They revealed two highly divergent molecular clades referred to as F and S, that do not conform to the morphological delineations of cyclotis and africana. A non‐negligible proportion of specimens of L. a. africana display haplotypes prevailing in forest elephant populations (clade F). The geographic distribution of clades and areas of their co‐occurrence support the hypothesis of incomplete isolation between forest and savannah African elephant populations, followed by recurrent interbreeding between the two forms. We state that the conclusions of prior studies resulted from insufficient character and/or geographic sampling. We conclude that there is no satisfying argument which can recognize two or more species of African elephants. We briefly comment on the meaning of such an attitude in a conservation viewpoint. © The Willi Hennig Society 2005.     

Movement patterns of forest elephants (Loxodonta cyclotis Matschie, 1900) in the Odzala-Kokoua National Park,Republic of Congo

African forest elephants (Loxodonta cyclotis Matschie, 1900) are ecological engineers that play a fundamental role in vegetation dynamics. The species is of immediate conservation concern, yet it is relatively understudied. To narrow this knowledge gap, we studied the drivers of daily movement patterns (linear displacements) of forest elephants—characterised by a set of geographical, meteorological and anthropogenic variables—in the Odzala-Kokoua National Park, Republic of Congo. Explicitly, we used conditional random forest to model and disentangle the main environmental factors governing the displacements of six forest elephants, fitted with GPS collars and tracked over 16 months. Results indicated that females moved further distances than males, while the presence of roads or human settlements disrupted elephant behaviour resulting in faster displacements. Forest elephants moved faster along watercourses and through forest with understory dominated by Marantaceae forests and bais, but moved slower in savannahs. Finally, flood-prone areas—described by elevation and accumulated precipitation—and higher temperatures prevented longer displacements. We expect these results to improve the knowledge on the species movements through different habitats, which would benefit its conservation management.     

Heard but not seen: an acoustic survey of the African forest elephant population at Kakum Conservation Area, Ghana

This study, designed to survey forest elephants ( Loxodonta africana cyclotis ) at Kakum Conservation Area, Ghana, is the first to apply acoustic methods to elephant abundance estimation and to compare results with independent survey estimates. Nine acoustic sensors gathered sound continuously for 38 days. Low-frequency calling rates have been established as useful elephant abundance indices at a Namibian watering hole and a central African forest clearing. In this study, we estimated elephant population size by applying an abundance index model and detection function developed in central Africa to data from simultaneous sampling periods on Kakum sensors. The sensor array recorded an average of 1.81 calls per 20-min sampling period from an effective detection area averaging 10.27 km². The resulting estimate of 294 elephants (95% CI: 259–329) falls within confidence bounds of recent dung-based surveys. An extended acoustic model, estimating the frequency with which elephants are silent when present, yields an estimate of 350 elephants (95% CI: 315–384). Acoustic survey confidence intervals are at least half as wide as those from dung-based surveys. This study demonstrates that acoustic surveying is a valuable tool for estimating elephant abundance, as well as for detecting other vocal species and anthropogenic noises that may be associated with poaching.     

Forest Elephants: Tree Planters of the Congo

The abundance of large vertebrates is rapidly declining, particularly in the tropics where over-hunting has left many forests structurally intact but devoid of large animals. An urgent question then, is whether these 'empty' forests can sustain their biodiversity without large vertebrates. Here we examine the role of forest elephant ( Loxodonta africana cyclotis ) seed dispersal in maintaining the community structure of trees in the Ndoki Forest, northern Congo. Analysis of 855 elephant dung piles suggested that forest elephants disperse more intact seeds than any other species or genus of large vertebrate in African forests, while GPS telemetry data showed that forest elephants regularly disperse seeds over unprecedented distances compared to other dispersers. Our analysis of the spatial distribution of trees from a sample of 5667 individuals showed that dispersal mechanism was tightly correlated with the scale of spatial aggregation. Increasing amounts of elephant seed dispersal was associated with decreasing aggregation. At distances of<200 m, trees whose seeds are dispersed only by elephants were less aggregated than the random expectation, suggesting Janzen–Connell effects on seed/seedling mortality. At the landscape scale, seed dispersal mode predicted the rate at which local tree community similarity decayed in space. Our results suggest that the loss of forest elephants (and other large-bodied dispersers) may lead to a wave of recruitment failure among animal-dispersed tree species, and favor regeneration of the species-poor abiotically dispersed guild of trees.     

Proboscidean mitogenomics: chronology and mode of elephant evolution using mastodon as outgroup

We have sequenced the complete mitochondrial genome of the extinct American mastodon (Mammut americanum) from an Alaskan fossil that is between 50,000 and 130,000 y old, extending the age range of genomic analyses by almost a complete glacial cycle. The sequence we obtained is substantially different from previously reported partial mastodon mitochondrial DNA sequences. By comparing those partial sequences to other proboscidean sequences, we conclude that we have obtained the first sequence of mastodon DNA ever reported. Using the sequence of the mastodon, which diverged 24–28 million years ago (mya) from the Elephantidae lineage, as an outgroup, we infer that the ancestors of African elephants diverged from the lineage leading to mammoths and Asian elephants approximately 7.6 mya and that mammoths and Asian elephants diverged approximately 6.7 mya. We also conclude that the nuclear genomes of the African savannah and forest elephants diverged approximately 4.0 mya, supporting the view that these two groups represent different species. Finally, we found the mitochondrial mutation rate of proboscideans to be roughly half of the rate in primates during at least the last 24 million years.     

Evolutionary and demographic processes shaping geographic patterns of genetic diversity in a keystone species,the African forest elephant (Loxodonta cyclotis)

The past processes that have shaped geographic patterns of genetic diversity may be difficult to infer from current patterns. However, in species with sex differences in dispersal, differing phylogeographic patterns between mitochondrial (mt) and nuclear (nu) DNA may provide contrasting insights into past events. Forest elephants (Loxodonta cyclotis) were impacted by climate and habitat change during the Pleistocene, which likely shaped phylogeographic patterns in mitochondrial (mt) DNA that have persisted due to limited female dispersal. By contrast, the nuclear (nu) DNA phylogeography of forest elephants in Central Africa has not been determined. We therefore examined the population structure of Central African forest elephants by genotyping 94 individuals from six localities at 21 microsatellite loci. Between forest elephants in western and eastern Congolian forests, there was only modest genetic differentiation, a pattern highly discordant with that of mtDNA. Nuclear genetic patterns are consistent with isolation by distance. Alternatively, male‐mediated gene flow may have reduced the previous regional differentiation in Central Africa suggested by mtDNA patterns, which likely reflect forest fragmentation during the Pleistocene. In species like elephants, male‐mediated gene flow erases the nuclear genetic signatures of past climate and habitat changes, but these continue to persist as patterns in mtDNA because females do not disperse. Conservation implications of these results are discussed.     

Roadless wilderness area determines forest elephant movements in the Congo Basin

A dramatic expansion of road building is underway in the Congo Basin fuelled by private enterprise, international aid, and government aspirations. Among the great wilderness areas on earth, the Congo Basin is outstanding for its high biodiversity, particularly mobile megafauna including forest elephants (Loxodonta africana cyclotis). The abundance of many mammal species in the Basin increases with distance from roads due to hunting pressure, but the impacts of road proliferation on the movements of individuals are unknown. We investigated the ranging behaviour of forest elephants in relation to roads and roadless wilderness by fitting GPS telemetry collars onto a sample of 28 forest elephants living in six priority conservation areas. We show that the size of roadless wilderness is a strong determinant of home range size in this species. Though our study sites included the largest wilderness areas in central African forests, none of 4 home range metrics we calculated, including core area, tended toward an asymptote with increasing wilderness size, suggesting that uninhibited ranging in forest elephants no longer exists. Furthermore we show that roads outside protected areas which are not protected from hunting are a formidable barrier to movement while roads inside protected areas are not. Only 1 elephant from our sample crossed an unprotected road. During crossings her mean speed increased 14-fold compared to normal movements. Forest elephants are increasingly confined and constrained by roads across the Congo Basin which is reducing effective habitat availability and isolating populations, significantly threatening long term conservation efforts. If the current road development trajectory continues, forest wildernesses and the forest elephants they contain will collapse.     

PHYLOGEOGRAPHY OF THE ASIAN ELEPHANT (ELEPHAS MAXIMUS) BASED ON MITOCHONDRIAL DNA

Abstract Populations of the Asian elephant (Elephas maximus) have been reduced in size and become highly fragmented during the past 3000 to 4000 years. Historical records reveal elephant dispersal by humans via trade and war. How have these anthropogenic impacts affected genetic variation and structure of Asian elephant populations? We sequenced mitochondrial DNA (mtDNA) to assay genetic variation and phylogeography across much of the Asian elephant's range. Initially we compare cytochrome b sequences (cyt b) between nine Asian and five African elephants and use the fossil‐based age of their separation (～5 million years ago) to obtain a rate of about 0.013 (95% CI = 0.011–0.018) corrected sequence divergence per million years. We also assess variation in part of the mtDNA control region (CR) and adjacent tRNA genes in 57 Asian elephants from seven countries (Sri Lanka, India, Nepal, Myanmar, Thailand, Malaysia, and Indonesia). Asian elephants have typical levels of mtDNA variation, and coalescence analyses suggest their populations were growing in the late Pleistocene. Reconstructed phylogenies reveal two major clades (A and B) differing on average by HKY85/Γ‐corrected distances of 0.020 for cyt b and 0.050 for the CR segment (corresponding to a coalescence time based on our cyt b rate of ～1.2 million years). Individuals of both major clades exist in all locations but Indonesia and Malaysia. Most elephants from Malaysia and all from Indonesia are in well‐supported, basal clades within clade A, thus supporting their status as evolutionarily significant units (ESUs). The proportion of clade A individuals decreases to the north, which could result from retention and subsequent loss of ancient lineages in long‐term stable populations or, perhaps more likely, via recent mixing of two expanding populations that were isolated in the mid‐Pleistocene. The distribution of clade A individuals appears to have been impacted by human trade in elephants among Myanmar, Sri Lanka, and India, and the subspecies and ESU statuses of Sri Lankan elephants are not supported by molecular data.     

Origin and phylogeography of African savannah elephants (<Emphasis Type="Italic">Loxodonta africana</Emphasis>) in Kruger and nearby parks in southern Africa

African savannah elephants (Loxodonta africana) occur in fragmented and isolated populations across southern Africa. Transfrontier conservation efforts aim at preventing the negative effects of population fragmentation by maintaining and restoring linkages between protected areas. We sought to identify genetic linkages by comparing the elephants in Kruger National Park (South Africa) to populations in nearby countries (Botswana, Mozambique, Zambia and Zimbabwe). We used a 446 base pair mitochondrial DNA (mtDNA) control region fragment (141 individuals) and 9 nuclear DNA (nDNA) microsatellite markers (69 individuals) to investigate phylogenetic relationships and gene flow among elephant populations. The mtDNA and nDNA phylogeographic patterns were incongruent, with mtDNA patterns likely reflecting the effects of ancient female migrations, with patterns persisting due to female philopatry, and nDNA patterns likely reflecting male-mediated dispersal. Kruger elephant heterozygosity and differentiation were examined, and were not consistent with genetic isolation, a depleted gene pool or a strong founder effect. Mitochondrial DNA geographic patterns suggested that the Kruger population was founded by elephants from areas both north and south of Kruger, or has been augmented through migration from more than one geographic source. We discuss our findings in light of the need for conservation initiatives that aim at maintaining or restoring connectivity among populations. Such initiatives may provide a sustainable, self-regulating management approach for elephants in southern Africa while maintaining genetic diversity within and gene flow between Kruger and nearby regions.     

Forest elephant mitochondrial genomes reveal that elephantid diversification in Africa tracked climate transitions

Among elephants, the phylogeographic patterns of mitochondrial (mt) and nuclear markers are often incongruent. One hypothesis attributes this to sex differences in dispersal and in the variance of reproductive success. We tested this hypothesis by examining the coalescent dates of genetic markers within elephantid lineages, predicting that lower dispersal and lower variance in reproductive success among females would have increased mtDNA relative to nuclear coalescent dates. We sequenced the mitochondrial genomes of two forest elephants, aligning them to mitogenomes of African savanna and Asian elephants, and of woolly mammoths, including the most divergent mitogenomes within each lineage. Using fossil calibrations, the divergence between African elephant F and S clade mitochondrial genomes (originating in forest and savanna elephant lineages, respectively) was estimated as 5.5 Ma. We estimated that the (African) ancestor of the mammoth and Asian elephant lineages diverged 6.0 Ma, indicating that four elephantid lineages had differentiated in Africa by the Miocene–Pliocene transition, concurrent with drier climates. The coalescent date for forest elephant mtDNAs was c. 2.4 Ma, suggesting that the decrease in tropical forest cover during the Pleistocene isolated distinct African forest elephant lineages. For all elephantid lineages, the ratio of mtDNA to nuclear coalescent dates was much greater than 0.25. This is consistent with the expectation that sex differences in dispersal and in variance of reproductive success would have increased the effective population size of mtDNA relative to nuclear markers in elephantids, contributing to the persistence of incongruent mtDNA phylogeographic patterns.     

From left to right all through the night: Characteristics of lying rest in zoo elephants

Despite increased research during the past years, many characteristics of resting behavior in elephants are still unknown. For example, there is only limited data suggesting elephants express longer lying bouts and increased total nightly lying durations on soft substrates as compared to hard surfaces. Additionally, it has not been investigated how frequently elephants change body sides between lying bouts. Here we present these characteristics based on observations of nighttime lying behavior in 10 zoo elephants (5 African Loxodonta africana and 5 Asian Elephas maximus elephants) living in five different European facilities. We found that elephants housed on soft substrates have significantly increased total lying durations per night and longer average lying bouts. Furthermore, at 70%−85% of all bouts, a consistently higher frequency of side change between lying bouts occurred on soft substrates, leading to an overall equal laterality in resting behavior. Deviations from this pattern became evident in elephants living on nonsand flooring or/and in nondominant individuals of nonfamily groups, respectively. Based on our findings, we consider elephants to normally have several lying bouts per night with frequent side changes, given an appropriate substrate and healthy social environment. We encourage elephant-keeping facilities to monitor these characteristics in their elephants' nighttime behavior to determine opportunities for further improvements and detect alterations putatively indicating social or health problems in individual elephants at an early stage.