Sourcing Elephant Ivory from a Sixteenth-Century Portuguese Shipwreck

The shipwreck and its contents are well preserved (Figure 1), and we were able to extract DNA successfully from 44 of 62 (71%) tusks available. Using targeted amplicon sequencing on the shipwreck ivory DNA, we examined three short unlinked chromosome segments for single-nucleotide polymorphisms (SNPs) fixed between African savanna (Loxodonta africana) and African forest (L. cyclotis) elephants. The character states detected were always diagnostic of forest and never of savanna elephants (Table S1). This indicated that the ivory derived from forest elephants, which historically ranged across the entire Guinean and Congolian tropical forest blocks of West and Central Africa (Figure 2).

Female elephants remain with natal social groups (“herds”) in relatively restricted geographic ranges. Thus, mitochondrial (mt)DNA, which is maternally transmitted, can often identify the geographic provenance of elephants., A ∼436-bp region of D-loop mtDNA was amplified for 44 shipwreck ivory samples (Table S2). African elephant mtDNA is geographically structured into 8 subclades (Figure 3A). In a network with published sequences representative of these 8 subclades (Figure 3B; Table S2), 23 of the 44 ivory samples grouped with the Western mtDNA subclade, carried only by elephants from West Africa. The other 21 grouped within the West-Central subclade, carried by elephants in both West and Central Africa (Figure 3A). For 17 of 44 shipwreck ivory samples, complete mitogenomes were assembled (see STAR Methods). Each of the 17 carried a distinct mitogenome haplotype, and when compared to 11 published mitogenomes (Table S2), the shipwreck ivory grouped with elephants from West Africa (Figure 3D).

Each mtDNA subclade consists of distinctive sequence haplotypes, many with very limited geographic distributions. A 336-bp segment of the mtDNA D-loop control region of the shipwreck ivory was compared to the 37 published mtDNA sequences within the West-Central (n = 29) and Western (n = 8) mtDNA subclades (Table S2; Figure 3C). The ivory exactly matched 3 haplotypes reported only from West Africa, with one exact match to a haplotype found in both West and Central Africa and no matches to haplotypes carried exclusively by Central African elephants (Figure 3C). Additionally, 16 novel geographically referenced elephant samples collected across Africa in the late twentieth century were sequenced for the same mtDNA region as the ivory (Table S2). The shipwreck ivory haplotypes grouped with haplotypes of the newly sequenced elephants that were from West Africa (Figure S1).

The shipwreck ivory samples lacked haplotypes from Northern-savanna, Savanna-wide, and Southeast-savanna subclades, present only among savanna elephants (see Figure 3A), consistent with the Bom Jesus ivory being exclusively from forest elephants. The shipwreck ivory carried no haplotypes from the North-Central, East-Central, or South-Central subclades, which would almost certainly have been present among tusks harvested in Central Africa (Figure 3A). Thus, all lines of evidence consistently indicated a West African forest elephant origin for all of the shipwreck ivory. The West African origin for the tusks also accords with historical records. Raw and carved ivories were exported from the Atlantic coast of Africa to Portugal from the mid-fifteenth century., Of three major regions believed to export ivory to Portugal,,, our genetic findings were consistent with an origin in Senegambia or the Gulf of Guinea (Figure 2) but rule out the Loango coast in Central Africa.

To further examine the geographic source(s) of the ivory, carbon and nitrogen stable isotope ratios were measured (successfully for 97 of 100 tusks available) to determine the diets and habitats of the elephants. Stable carbon isotope ratios measured in ivory originate from the plant food the elephant consumed, which in turn is an indication of their habitats. Ivory has more positive δ13C values than plant food due to further partitioning of carbon isotopes during digestion and tissue synthesis, with a diet-tissue enrichment of ∼5.5‰ reported in modern and fossil proboscideans. Published δ13Ccollagen for wild African elephants range from −27‰ for pure C3 feeders in deep rainforests to −11‰ for savanna elephants consuming substantial quantities of C4 grass (Figure 4). In the shipwreck ivory collagen, δ13C values averaged −20.4‰ ± 1.2‰, with a range from −22.2‰ to −17.1‰ (Figure 4; Table S3). Shipwreck ivory δ13C values are more positive than those of modern elephants from Cameroon, Democratic Republic of Congo, and Liberia, which have 13C-depleted values reflecting pure C3 diets in deep forest/rainforest habitats with continuous tree canopy., , ,  Shipwreck samples with the most positive δ13C values fall within the range of modern comparative samples from open or shrub savanna environments, where elephants consume substantial proportions of C4 grasses., , 

Variation of 15N in elephants is driven primarily by nitrogen cycling in the soil, which is strongly influenced by moisture availability., , ,  Published δ15Ncollagen values for wild African elephants range from 2‰ in moist areas to 17‰ in arid areas.,,,, , , ,  In the shipwreck ivory, δ15N values averaged 6.8‰ ± 0.8‰, with a range from 4.8‰ to 9.0‰ (Figure 4). Such values typically derive from mesic terrestrial environments, falling in the middle of the range of δ15N values documented for African elephants and encompassing the values for elephants living in habitats in Angola, Benin, Burkina Faso, Chad, Central African Republic, and Niger. The shipwreck ivory δ15N is lower than those (above 10‰) of elephants in open, arid grassland savanna environments (Angola, Namibia, and South Africa) or living in dense forests (Democratic Republic of Congo).,

Values for the shipwreck ivory extend over 46% of the total variation in δ13C and 49% of the total variation in δ15N across African elephants (Figure 4)., The broad range suggests that the ivory was sourced from elephants from different habitats with different rainfall regimes and vegetation, although not from deep rainforests or arid environments. For four tusks, we took additional incremental samples to examine within-individual dietary variation over time (Figure S2). The δ13C values from the incremental sampling for all four of these tusks are indicative of elephants that lived in mixed savanna and woodland habitats during the time the tusks were growing. The longest sequence derives from tusk B6082: δ13C and δ15N values show a cyclical pattern of increasing and decreasing over a series of 22 growth increments, indicative of switching between C3 browse and C4 graze in the dry and wet seasons, respectively.,,, The range of δ13C values (6‰) in this tusk is consistent with a habitat outside of the deep tropical forest, for which seasonal variation in δ13C is typically ≤2‰.

In West Africa, dense tropical lowland rainforests of the Guinean forest block are surrounded by dry forest and thicket and a mosaic of forest and savanna habitats (Figure 2). Based on combined genetic and isotope analyses, the elephants hunted for the ivory cargo of the Bom Jesus originated from West African forest elephants in habitats outside the Guinean forest block. In 1482, the Portuguese built a fort at an established trading post in West Africa, São Jorge de Mina, or Elmina, on the south-western edge of the Dahomey (or Benin) Gap., This is a long-established area of savanna and drier-type forest vegetation that provided an important corridor for transporting goods (including ivory) from the interior to the coast, thus avoiding travel through dense forest. Settlements near the Dahomey Gap then expanded as Elmina became an important entrepôt.

Due to the difficulty of maneuvering large long-distance trading vessels and the dangers of sailing close to the shore, outgoing ships on the India route typically did not tack along the West African coastline but would sail from Portugal southwest across the Atlantic and then southeast on the trade winds., Ivory from West Africa was frequently shipped to the islands of Cape Verde and São Tomé to be counted, weighed, and sent via smaller vessels to Casa da Índia in Lisbon, the central clearing house for African and Indian imports to Portugal.,,, Centralized loading of outgoing long-distance trading vessels (naus) in Lisbon enabled tight control of the valuable cargo. Although the Great Lisbon earthquake and fire of 1755 destroyed many of the archives of the Casa da Índia, the consolidation of ivory from different localities within West Africa accords with our findings: the range of isotope values suggests multiple different habitats, as does the presence of 17 distinct mitogenome haplotypes, indicating that the Bom Jesus ivory derived from at least 17 different “herds” of elephants.

In West Africa, the historic range of savanna elephants was likely continuous across the Sahelian/Sudanian savanna habitat belts north of the Guinean forest block. The current distributional range of forest elephants includes habitats both in tropical forest and in nearby savannas., , , Before our analyses, the recent distribution of forest elephants in West Africa outside of tropical forest habitats could be attributed to decimation of savanna elephants in West Africa in the 19th and 20th centuries., Our combined genetic and isotope results instead indicate that utilization of savanna habitats by forest elephants in West Africa preceded the removal of savanna elephants and dates back to at least the sixteenth century.

The Bom Jesus tusks are of varying length and size (from 2 to 33 kg), and the elephants may have been hunted indiscriminately, both males and females, young and old alike., Among the mtDNA haplotypes identified from the sixteenth-century tusks, only four have been reported among contemporary populations (Figure 3C), likely reflecting the impact of the ivory trade and reduction of historic elephant range by at least 93% in West Africa. Decrease in population size and genetic diversity are associated with negative conservation outcomes, such as expression of deleterious alleles, reduced reproductive fitness, and increased risk of population extirpation. Our geo-referenced sequences from late-twentieth-century elephants in the University of Cape Town (UCT) collection, and the newly sequenced shipwreck ivory sequences, add substantially to the previously small body of isotopic and genetic information for West African elephants, with the potential to aid in the sourcing of confiscated illegal ivory., Improving the ability to trace poached ivory can help guide optimal allocation of scarce law enforcement resources.

The Bom Jesus was one of 80 vessels that sailed from Lisbon on the India route between 1531 and 1540. This was one of the most strategic and lucrative commercial routes of the time, linking to established trading networks between Africa and Asia., The large number of tusks recovered from the Bom Jesus is evidence of ivory acquisition and circulation driving the formative stages of globalization. With a resolution not possible using any single approach, our interdisciplinary methodologies revealed the long-term genetic diversity and habitat use of the African forest elephant, helpful for conserving this iconic species. To refine the sourcing of archeological and historical ivory, future work can utilize the combination of genetic and isotope methods presented here and additional approaches in both fields as they are further developed.

Unworked elephant tusks, ivory working debris, and finished objects made from ivory have been recovered from numerous archaeological contexts worldwide, including, but not limited to, shipwrecks with ivory cargo reported from the Mediterranean Sea and the Atlantic, Pacific, and Indian Oceans., , , ,  Our methods are applicable to the vast collections of historic and archaeological ivories in museums across the globe., ,  Analyzing historic and archaeological ivories affords a window into human-animal relationships across thousands of years and can reveal the formative and changing patterns of exchange between people who lived oceans apart.,, , , ,  Our study on the largest archaeological cargo of African ivory ever found provides a framework for examining one of the world’s most important raw materials throughout human history.