Genetic diversity and population structure of the endangered marsupial Sarcophilus harrisii (Tasmanian devil)

Abstract

The Tasmanian devil (Sarcophilus harrisii) is threatened with extinction because of a contagious cancer known as Devil Facial Tumor Disease. The inability to mount an immune response and to reject these tumors might be caused by a lack of genetic diversity within a dwindling population. Here we report a whole-genome analysis of two animals originating from extreme northwest and southeast Tasmania, the maximal geographic spread, together with the genome from a tumor taken from one of them. A 3.3-Gb de novo assembly of the sequence data from two complementary next-generation sequencing platforms was used to identify 1 million polymorphic genomic positions, roughly one-quarter of the number observed between two genetically distant human genomes. Analysis of 14 complete mitochondrial genomes from current and museum specimens, as well as mitochondrial and nuclear SNP markers in 175 animals, suggests that the observed low genetic diversity in today's population preceded the Devil Facial Tumor Disease disease outbreak by at least 100 y. Using a genetically characterized breeding stock based on the genome sequence will enable preservation of the extant genetic diversity in future Tasmanian devil populations.

Fig. 1.

Sequence coverage depth used for genetic variant detection. The coverage was calculated for Illumina sequences used for our three specimens in SNP calling against a de novo assembled reference sequence (14x coverage 454/Roche and Illumina hybrid assembly), and does not include potential PCR duplicates and secondary alignments. The y axis indicates the fraction of the non-N bases in the reference sequence that have a particular coverage. Vertical lines on the x axis indicate average coverage for the three samples.

Fig. 2.

Genetic diversity of Sarcophilus. (A) The numbers of heterozygous sites in Cedric and Spirit (in millions), and the number shared between them, compared with two human pairs (the only other vertebrate species for which strictly comparable data are available). Sarcophilus has far fewer such sites. In addition, a much higher fraction is shared between individuals, indicating less population stratification than in humans (see SI Appendix). (B) Mitochondrial diversity covering the last 100 y. Locations of single nucleotide variations (neglecting the hypervariable region) are indicated as vertical lines in the seven modern and six museum specimens relative to the eastern-derived animal, Spirit. Diversity ranges from the geographically most western animal (Cedric) to the most distant eastern animal (Spirit). (C) Average numbers of mitochondrial genome differences between pairs of individuals, ignoring hypervariable regions. Species designated by the 2008 IUCN Red List of Threatened Species as “endangered” or “critically endangered” are indicated in red, and extinct species are in black. Species and populations in blue are thriving. ^†Species represented by only two sequences. *Whales are averaged over five species. Woolly mammoths are divided into two mitochondrial clades (30). The gorillas may be from separate subspecies, Gorilla gorilla and Gorilla beringei. It is apparent that mitochondrial diversity is not the only factor affecting species endangerment; habitat loss and other factors are often critical.

Fig. 3.

Population structure of Sarcophilus. (A) Mitochondrial diversity map of Tasmania for animals from 17 locations, including some now in a mainland captive breeding program. Pie charts depict the location and size (number of animals) of individual populations. Identifiers for modern animals included in the complete mitochondrial sequencing are indicated in red. Four major mitochondrial haplogroups (A, B, C, and E) were identified. A fifth minor haplogroup, D, was found predominantly in the single off-shore captive breeding population. (B) Principal-components analysis scatter plot for 702 nuclear SNPs genotyped for 87 Tasmanian devils from 12 geographical locations reveals population substructure and diversity. Two core populations of low genetic diversity are found in the northwest and Bronte Park central regions of Tasmania. Although there is clustering of the eastern populations, each adds a unique subpopulation to this broad cluster. Two-letter codes can be inferred from A. (C) Partitions of Tasmania into regions where equal numbers of individuals for a captive breeding program should be chosen, based on our data.