Conservation genetics of the white-bellied pangolin in West Africa: A story of lineage admixture, declining demography, and wide sourcing by urban bushmeat markets.

During the last 40 years, the volumes of African pangolins feeding the illegal wildlife trade have dramatically increased. We conducted a conservation genetics survey of the most traded African species, the white bellied pangolin (WBP; Phataginus tricuspis), across three West African countries including Guinea, Côte d'Ivoire, and Ghana. Our study combining mitochondrial DNA sequencing and microsatellite genotyping is the first to reveal a wide pattern of admixture between two of the six mitochondrial lineages as previously delimited within WBP. We found a signature of isolation by distance but a lack of population genetic structuring, supporting the idea that WBP may have underestimated dispersal abilities. Levels of genetic diversity were low in West African lineages (WAfr and Gha) compared to Central Africa, reinforcing the picture of genetic pauperization shared by West African WBP. We observed a 85%-98% decline in the effective population size of WBP occurring c. 3200 to 400 ya, with current numbers (520-590) at the lower end of the conservative thresholds for minimum viable population size. The microsatellite markers were powerful enough to differentiate between individuals and identify replicated samples, confirming the utility of this approach in tracing the pangolin trade. Genetic diversity estimates confirmed that Yopougon, the main bushmeat market from Abidjan (Côte d'Ivoire), was fed by a large trade network as confirmed by vendors reporting 10 different sources situated 62-459 km away from the market. We conclude that WBP distributed in the Upper Guinean Block should be considered a single management unit of high conservation concern, as impacted by genetic diversity erosion, drastic decline in effective population size, and wide range sourcing for feeding urban bushmeat markets. Given the genetic admixture pattern detected within WBP from West Africa, we advocate for a multi-locus strategy to trace the international trade of the species.

Au cours des 40 dernières années, les volumes de pangolins africains alimentant le commerce illégal d'espèces sauvages ont augmenté de façon spectaculaire. Nous avons mené une étude de génétique de la conservation sur l'espèce africaine la plus braconnée, le pangolin à ventre blanc (PVB; Phataginus tricuspis), à travers trois pays d'Afrique de l'Ouest incluant la Guinée, la Côte d'Ivoire et le Ghana. Notre étude combinant le séquençage de l'ADN mitochondrial et le génotypage par microsatellites est la première à révéler un patron généralisé d'admixture entre deux des six lignées mitochondriales précédemment délimitées au sein du PVB. Nous mettons en évidence une signature d'isolement par la distance mais un manque de structuration génétique des populations, soutenant l'idée que le PVB a des capacités de dispersion jusque­là sous­estimées. Les niveaux de diversité génétique observés dans les lignées d'Afrique de l'Ouest (WAfr et Gha) sont faibles par rapport à ceux observés en Afrique Centrale, confirmant la paupérisation génétique des populations de PVB en Afrique de l'Ouest. Nous avons observé un déclin de 85 à 98% de la taille efficace de la population ouest­africaine entre 3200 et 400 ans, les estimations actuelles (520­590) se situant à l'extrémité inférieure des seuils minimums de viabilité. Les marqueurs microsatellites utilisés sont suffisamment puissants pour différencier les individus et identifier les doublons d'échantillons, confirmant l'utilité de ces marqueurs pour tracer le commerce des pangolins. Nos estimations de diversité génétique ont confirmé que Yopougon, le principal marché de viande de brousse d'Abidjan (Côte d'Ivoire), était alimenté par un vaste réseau commercial, comme le confirment les vendeurs signalant 10 sources différentes situées à 62­459 km du marché. Nous concluons que les PVB présents dans le bloc forestier de Haute Guinée devraient être considérés comme une seule unité de gestion à fort intérêt de conservation, étant soumis à une érosion de sa diversité génétique, un déclin drastique de la taille efficace de sa population et un approvisionnement à grande échelle des marchés urbains de viande de brousse. Compte tenu du schéma d'admixture génétique détecté chez les PVB d'Afrique de l'Ouest, nous préconisons une stratégie multi­locus pour tracer le commerce international de l'espèce.

Rediscovery, range extension, phylogenetic relationships and updated diagnosis of the Ornate Long-tailed Lizard Latastia ornata Monard, 1940 (Squamata: Lacertidae).

The lacertid Latastia ornata was known to date only by its holotype collected in 1938 in Bafatá, central Guinea-Bissau. We report new specimens and localities from Guinea-Conakry, a new country record and major range extension of 700 km SE of the type-locality. We provide an updated diagnosis of the species, including the first genetic and osteological data, and confirm that Latastia ornata is closely related to, but distinct from, L. longicaudata based on external morphology, cranial osteology, DNA data and zoogeography.

The Complete Phylogeny of Pangolins: Scaling Up Resources for the Molecular Tracing of the Most Trafficked Mammals on Earth.

Pangolins, considered the most-trafficked mammals on Earth, are rapidly heading to extinction. Eight extant species of these African and Asian scale-bodied anteaters are commonly recognized, but their evolutionary relationships remain largely unexplored. Here, we present the most comprehensive phylogenetic assessment of pangolins, based on genetic variation of complete mitogenomes and 9 nuclear genes. We confirm deep divergence among Asian and African pangolins occurring not later than the Oligocene-Miocene boundary ca. 23 million years ago (Ma) (95% HPD = 18.7-27.2), limited fossil evidence suggesting dispersals from Europe. We recognize 3 genera including Manis (Asian pangolins), Smutsia (large African pangolins), and Phataginus (small African pangolins), which first diversified in the Middle-Upper Miocene (9.8-13.3 Ma) through a period of gradual cooling coinciding with a worldwide taxonomic diversification among mammals. Based on large mitogenomic distances among the 3 genera (18.3-22.8%) and numerous (18) morphological traits unique to Phataginus, we propose the subfamily Phatagininae subfam. nov. to designate small African pangolins. In contrast with the morphological-based literature, our results establish that the thick-tailed pangolin (Manis crassicaudata) is sister-species of the Sunda (Manis javanica) and Palawan (Manis culionensis) pangolins. Mitogenomic phylogenetic delineations supported additional pangolin species subdivisions (n = 13), including 6 African common pangolin (Phataginus tricuspis) lineages, but these patterns were not fully supported by our multi-locus approach. Finally, we identified more than 5000 informative mitogenomic sites and diagnostic variation from 5 nuclear genes among all species and lineages of pangolins, providing an important resource for further research and for effectively tracing the worldwide pangolin trade.

Phylogeography of the heavily poached African common pangolin (Pholidota, Manis tricuspis) reveals six cryptic lineages as traceable signatures of Pleistocene diversification.

Knowledge on faunal diversification in African rainforests remains scarce. We used phylogeography to assess (i) the role of Pleistocene climatic oscillations in the diversification of the African common pangolin (Manis tricuspis) and (ii) the utility of our multilocus approach for taxonomic delineation and trade tracing of this heavily poached species. We sequenced 101 individuals for two mitochondrial DNA (mtDNA), two nuclear DNA and one Y-borne gene fragments (totalizing 2602 bp). We used a time-calibrated, Bayesian inference phylogenetic framework and conducted character-based, genetic and phylogenetic delineation of species hypotheses within African common pangolins. We identified six geographic lineages partitioned into western Africa, Ghana, the Dahomey Gap, western central Africa, Gabon and central Africa, all diverging during the Middle to Late Pleistocene. MtDNA (cytochrome b + control region) was the sole locus to provide diagnostic characters for each of the six lineages. Tree-based Bayesian delimitation methods using single- and multilocus approaches gave high support for 'species' level recognition of the six African common pangolin lineages. Although the diversification of African common pangolins occurred during Pleistocene cyclical glaciations, causative correlation with traditional rainforest refugia and riverine barriers in Africa was not straightforward. We conclude on the existence of six cryptic lineages within African common pangolins, which might be of major relevance for future conservation strategies. The high discriminative power of the mtDNA markers used in this study should allow an efficient molecular tracing of the regional origin of African common pangolin seizures.

Bushmeat genetics: setting up a reference framework for the DNA typing of African forest bushmeat.

The bushmeat trade in tropical Africa represents illegal, unsustainable off-takes of millions of tons of wild game - mostly mammals - per year. We sequenced four mitochondrial gene fragments (cyt b, COI, 12S, 16S) in >300 bushmeat items representing nine mammalian orders and 59 morphological species from five western and central African countries (Guinea, Ghana, Nigeria, Cameroon and Equatorial Guinea). Our objectives were to assess the efficiency of cross-species PCR amplification and to evaluate the usefulness of our multilocus approach for reliable bushmeat species identification. We provide a straightforward amplification protocol using a single 'universal' primer pair per gene that generally yielded >90% PCR success rates across orders and was robust to different types of meat preprocessing and DNA extraction protocols. For taxonomic identification, we set up a decision pipeline combining similarity- and tree-based approaches with an assessment of taxonomic expertise and coverage of the GENBANK database. Our multilocus approach permitted us to: (i) adjust for existing taxonomic gaps in GENBANK databases, (ii) assign to the species level 67% of the morphological species hypotheses and (iii) successfully identify samples with uncertain taxonomic attribution (preprocessed carcasses and cryptic lineages). High levels of genetic polymorphism across genes and taxa, together with the excellent resolution observed among species-level clusters (neighbour-joining trees and Klee diagrams) advocate the usefulness of our markers for bushmeat DNA typing. We formalize our DNA typing decision pipeline through an expert-curated query database - DNA BUSHMEAT - that shall permit the automated identification of African forest bushmeat items.

Reviving the African wolf Canis lupus lupaster in North and West Africa: a mitochondrial lineage ranging more than 6,000 km wide.

The recent discovery of a lineage of gray wolf in North-East Africa suggests the presence of a cryptic canid on the continent, the African wolf Canis lupus lupaster. We analyzed the mtDNA diversity (cytochrome b and control region) of a series of African Canis including wolf-like animals from North and West Africa. Our objectives were to assess the actual range of C. l. lupaster, to further estimate the genetic characteristics and demographic history of its lineage, and to question its taxonomic delineation from the golden jackal C. aureus, with which it has been considered synonymous. We confirmed the existence of four distinct lineages within the gray wolf, including C. lupus/familiaris (Holarctic wolves and dogs), C. l. pallipes, C. l. chanco and C. l. lupaster. Taxonomic assignment procedures identified wolf-like individuals from Algeria, Mali and Senegal, as belonging to C. l. lupaster, expanding its known distribution c. 6,000 km to the west. We estimated that the African wolf lineage (i) had the highest level of genetic diversity within C. lupus, (ii) coalesced during the Late Pleistocene, contemporaneously with Holarctic wolves and dogs, and (iii) had an effective population size of c. 80,000 females. Our results suggest that the African wolf is a relatively ancient gray wolf lineage with a fairly large, past effective population size, as also suggested by the Pleistocene fossil record. Unique field observations in Senegal allowed us to provide a morphological and behavioral diagnosis of the African wolf that clearly distinguished it from the sympatric golden jackal. However, the detection of C. l. lupaster mtDNA haplotypes in C. aureus from Senegal brings the delineation between the African wolf and the golden jackal into question. In terms of conservation, it appears urgent to further characterize the status of the African wolf with regard to the African golden jackal.