NEW YORK (GenomeWeb News) – In a study appearing online last night in the Proceedings of the National Academy of Sciences, researchers from Pennsylvania State University and elsewhere describe how they used whole nuclear and mitochondrial genome sequencing and genotyping to characterize genetic diversity and population structure in Tasmanian devils — information that they hope will aid conservation efforts for the animals, which have been devastated by a contagious cancer known as devil facial tumor disease (DFTD).
"Genomics, based on next-generation sequencing, can be applied for the preservation of endangered species," co-corresponding author Stephan Schuster, a biochemistry and molecular biology researcher at Penn State, told GenomeWeb Daily News. "The Tasmanian devil is our first showcase, where we tried to demonstrate the power of this approach."
The researchers sequenced the nuclear genomes of two Tasmanian devils with different susceptibilities to DFTD from opposite sides of the Australian island, along with the genome of a tumor sample from one of the animals. Coupled with mitochondrial genome data on modern-day devils and museum samples collected over more than 130 years, sequence variants identified in the newly sequenced genomes offered clues about the animals' population structure on the Australian island, as well as insights into the genetic diversity found in Tasmanian devils now and in the past.
For instance, the study suggest that while genetic diversity in Tasmanian devil populations is low, it has not declined over the past 130 years or more, even with the arrival of DFTD, which was first detected in wild populations 15 years ago.
Based on their findings so far, those involved in the study advocate the use of genetic markers to inform Tasmanian devil captive breeding programs — a strategy that they say has thus far been met with resistance by those running such programs.
"With this kind of [genetics informed breeding] approach, you can absolutely guarantee that the remaining genetic diversity is contained in even a much, much smaller breeding population than what you would need if you do the random approach," Schuster argued.
DFTD, first detected in 1996, is caused by an infectious cancer cell line that spreads from one Tasmanian devil to another by biting and other contact between the animals. The disease has cut wild devil populations by as much as 90 percent in some parts of Tasmania, fueling speculation that the animals' susceptibility to the disease might stem from low genetic diversity in Tasmanian devil populations.
Even so, the researchers explained, estimates of this genetic diversity have been largely based on estimates from targeted analyses of specific genes, including genes involved in major histocompatibility complex mediated immunity.
For the current study, the team sequenced the nuclear genomes of a two devils: a male dubbed "Cedric," from northwestern Tasmania, who had survived two rounds of DFTD infection in the lab, and a female from the southeast, named "Spirit," who had been infected in the wild and succumbed to the disease shortly afterwards. They also sequenced the DFTD genome using one of the tumor samples isolated from Spirit.
Schuster presented preliminary findings from the study at the Plant and Animal Genomes conference in San Diego early this year.
To sequence the genomes, the team first used the Roche 454 GS FLX Titanium and an experimental, early access Roche 454 GS FLX Titanium XL instrument, which generated reads up to 800 base pairs long, to come up with a 3.3 billion base draft Tasmanian devil reference genome.
They then did paired-end sequencing with the Illumina GA IIx to get 16.7 times coverage of Cedric's genome, 32.2 times coverage of Spirit's genome, and 19.7 times coverage of the tumor genome.
When they compared Cedric and Spirit's nuclear genomes to one another, researchers detected 1,057,507 SNPs — around 22 percent of the genetic diversity expected in humans if one were to compare someone from the African Bushman population with someone from Japan, they noted.
The sequenced tumor sample, meanwhile, contained nearly 119,000 SNPs not detected in genome sequences from Cedric or Spirit, including 110 tumor-specific alleles that the team subsequently validated by amplicon sequencing with the Ion Torrent semi-conductor sequencing platform.
The team got additional clues about the genetic diversity in Tasmanian devil populations by sequencing the mitochondrial genomes of seven present-day devils, a tumor sample from Spirit, and six Tasmanian devils from museum collections.
On average, the researchers found 13 SNPs between Tasmanian devils from northwestern and southeastern parts of Tasmania, Schuster explained.
But when they compared the mitochondrial genomes over time, the team saw similar levels of genetic diversity, even for samples stretching back as far as 1874.
"I think this is very important," Schuster explained. "It means that, so far, the facial tumor has not decimated or brought down the genetic diversity any further than what it was 130 years ago."
Analyses of the mitochondrial genomes also yielded 17 SNPs that appeared to be useful for genotyping the animals, while comparisons of the nuclear genomes spawned 96- and 1,536-SNP marker sets that could be assessed using custom Illumina Golden-Gate arrays.
Indeed, the researchers' nuclear and mitochondrial genotyping experiments on 168 wild and seven captive animals helped them to map Tasmanian devil genetic patterns and population structure across Tasmania.
"Our approach is to take two individuals with the maximal geographic distance, maximize the number of genetic differences you discover by sequencing the nuclear genome, pick your genetic markers, and come up with a genetic test that only costs you a few hundred dollars … and then go after hundreds of animals," Schuster said.
Moreover, with respect to the conservation efforts underway to try to protect Tasmanian devils, Schuster said genetic markers such as those identified in the current study could help preserve the genetic diversity that remains in Tasmanian devil populations. Consequently, while he conceded that genetic testing would be an added expense for conservation programs, he believes such informed breeding strategies could pay off in the long run.
Data generated in the study is being made available to other researchers. Additional information on the Tasmanian Devil Genome Project is available here.