University of Minnesota researchers have successfully mapped the complete genome of the endangered Przewalski’s horse. Once extinct in the wild, the species now has a population of around 2,000 animals thanks to conservation efforts.

The study, published in the journal G3, was led by Nicole Flack and Lauren Hughes, researchers at the College of Veterinary Medicine, along with Christopher Faulk, a professor in the College of Food, Agricultural and Natural Resource Sciences. University of Minnesota students contributed to the genome sequencing through Faulk’s animal science course. 

“The genome is the basic blueprint for an animal and tells us what makes a species unique and also tells us about the health of a population,” said Faulk. “My students worked together to produce the highest quality Przewalski’s horse genome in the world.”

Researchers can now use this as a tool to make accurate predictions about what gene mutations mean for Przewalski’s horse health and conservation.  

“Studying genes without a good reference is like doing a 3 billion-piece puzzle without the picture on the box,” said Flack. “Przewalski’s horse researchers studying mutations in an important gene need a good reference picture to compare their puzzle with.” 

Researchers used a blood sample from Varuschka, a 10-year-old Przewalski’s mare at the Minnesota Zoo, to construct a representative map of genes for the species. The zoo has long been active in Przewalski’s horse breeding and management, with over 50 foals born since the 1970s. 

“We were excited to partner with the University of Minnesota to preserve the genetic health of the species as their populations continue to recover, both in zoos and in the wild,” said Anne Rivas, doctor of veterinary medicine at the Minnesota Zoo. “We are thrilled to offer our community the opportunity to see the horse as the results of our conservation efforts.” 

The cutting-edge technology sequencing used to construct the genome uses a small machine about the size of a soda can. Its portability means this method could be adapted for further study of wild Przewalski’s horses in remote locations.

Future applications of the reference genome may include studying genes that help the horse adapt to environmental changes, identifying mutations associated with specific traits or diseases, and informing future breeding decisions to help improve upon genetic diversity. Given the extreme population bottleneck that occurred during the near-extinction of Przewalski’s horse, such understanding is crucial for continued breeding efforts.

Look at any list of the top 10 most aggressive animals, and you will undoubtedly find a mugshot of the North American wolverine. Although much smaller than most of the other animals accompanying it on such lists—such as the hippopotamus and the wild boar—this feisty member of the weasel family has been a protagonist in popular myths, even serving as inspiration for superhero characters and as mascots of sports teams. A wide range of cultural, social, economic, and psychological factors influence human-wolverine relations. In many Indigenous societies, the wolverine is a respected cultural keystone species and is often viewed as a trickster. Unfortunately, these solitary carnivores rely on cold temperatures and snowy environments for their reproduction and survival, so their future remains uncertain in a warming world.

A new study published in the August issue of G3: Genes|Genomes|Genetics provides geneticists and conservation biologists with a high-quality, chromosome-level genome assembly of the wolverine, including extensive annotation of genes involved in behavior and immune responses to pathogens. The authors’ goal goes far beyond the wolverine: they seek to provide similar high-quality assemblies for other species predicted to be impacted by increasing global temperatures. This means setting the benchmark for a workflow that offers the best compromise possible when balancing cost, time, simplicity, accuracy, and completeness for long-read assembly and genome annotation. 

“Our goal is to replace the existing short-read assemblies and increase the quality standards for new reference genomes in light of current sequencing technologies,” says lead author Si Lok.

Improving the standards of genome assembly in the current era

The DNA sequenced in the report comes from a 30-year-old tissue sample of a male wolverine specimen from the Kugluktak (Coppermine) region of Nunavut. Lok and colleagues use PacBio contiguous long-reads (CLR) mode, which typically provides maximal read length at a reasonable cost; however, it is prone to 15-20% pseudo-random errors. To mitigate such inaccuracies while maintaining the numerous benefits of this approach, the authors used a two-step workflow for genome assembly: 1) the uncorrected CLRs are assembled using Flye assembler, followed by a polishing regimen with high-quality Illumina short reads, and 2) the subsequent scaffolding of the assembly against assemblies of related family members.

“It took us nearly two years to optimize a workflow that produces a final genome assembly comprising well less than 1000 contigs—about 10–100 times better than those found in most genome reports—at a cost of under $10,000,” says Lok. The cost is going down all the time.

This new workflow leads to striking completeness and accuracy: 99.98% of the current BUSCO set of 9,226 genes used to assess assembly quality are complete at exon-level in the wolverine assembly, placing it in the top tier of assemblies produced from long-reads. Lok hopes that their report shows how cost-effective, accurate, and complete sequencing and assembly can be nowadays. “No future genome reports should be less than chromosome-level, given the current technologies.”

Conservation genomics meets wolverine behavior

The new article also provides the first full-length mitochondrial genome assembly for the North American wolverine, as well as a tabulation of potential microsatellite markers for the wolverine. Since monitoring population size and distribution, reproductive success, and gene flow in wild populations often relies on analyses of mitochondrial DNA and microsatellites, the authors hope to provide a resource for developing these and other species-specific genomic markers.

In addition, Lok and colleagues annotated genes whose orthologs have been associated with aggressive traits in other organisms—an adaptation to drive competition for food and mates— and the key components of innate immune responses. “Environmental disruptions from climate change will increase vulnerabilities to new pathogens,” says Lok.

“We are in the process of reporting genomes for other species predicted to be heavily affected by climate change in efforts to support their conservation and ecological relationships, such as that of the Canada lynx and the snowshoe hare,” says Lok. He wishes this report to set a minimum standard of quality for future genome reports and resources for conservation biologists.

Efforts to diversify an inbred population must take into account the genetic backgrounds of the founders.

The nocturnal flightless parrot known as the kākāpō was once abundant throughout New Zealand. But after the introduction of mammalian predators, the species all but disappeared. Today, every living kākāpō is descended from a tiny handful of island survivors and a single male from the mainland. The entire population of 201 birds is closely watched over by conservationists on a few predator-free island refuges, where they hope the hefty green parrots will continue to breed.

But because their numbers dropped into the double digits, the kākāpō face a genetic bottleneck. Inbreeding can increase rates of genetic disease and contribute to poor health among the population. Geneticists studying the remaining kākāpō have quantified the amount of inbreeding among the birds as one way to understand the health of the species. Their results, reported in a new paper in G3: Genes|Genomes|Genetics, suggest that inbreeding isn’t necessarily hurting chicks’ chances of survival and that introducing additional genetic diversity may not always have the intended effect.

Saved from extinction

“Kākāpō have a really interesting natural history,” says Yasmin Foster, a graduate student at the University of Otago and the study’s lead author. “They were functionally extinct, but then a small population was found on an island in the south of New Zealand.”

“Functionally extinct” in this case meant no more females could be found on the New Zealand mainland; only a few males remained. In 1977, about 50 kākāpō were discovered living on Stewart Island, a large island about 19 miles south of the mainland. Predators such as feral cats roamed Stewart Island, however, so in 1982 conservationists began relocating the birds to several smaller, mammal-free outlying islands.

By then, only one male kākāpō remained on the mainland, and he was taken to a predator-free island refuge along with the Stewart Island population. The Stewart Island kākāpō had diverged from the mainland population around 10,000 years ago, giving the two groups time to develop distinct genetic profiles. Introducing the mainland bird was meant to help boost genetic diversity among a new generation of chicks.

Now, the population has grown to 201 individuals. To help inform conservation strategies, Foster and her colleagues set out to document the amount of inbreeding in the colony. Creating a large, multigenerational pedigree of the wild kākāpō wouldn’t work for a founder population of this type, so the researchers turned to DNA sequence analysis. Thanks to the availability of a high-quality kākāpō reference genome, the team could genotype the birds using genome-wide mapping of single nucleotide polymorphisms (SNPs).

Comparing measures of inbreeding

“We had this unique founding population with 50 Stewart Island birds and one mainland male,” says Foster. “From the inbreeding metrics I looked at, we found that they’re both inbred, but in different ways.” She says the study raises an interesting point about how combining two inbred populations in an attempt to increase genetic diversity can actually introduce more deleterious alleles.

Comparing multiple inbreeding metrics helped Foster get a robust view of the birds’ genome, and also to evaluate the accuracy of each one. “Some people just use one or another, but what I found was that some of them give a different story,” she says.

She started by calculating the coefficient of inbreeding, or F_H, which is the probability of an individual inheriting two copies of the same allele from the same ancestor on both sides. She compared this with a newer method of measurement, called “runs of homozygosity” or F_ROH, which looks for long sections of the genome where both copies are the same.

A third method, using pairwise analysis to construct a genomic-relatedness matrix, turned out to be the weakest of the metrics, Foster says. “Other people have found that when you have a small group of individuals that are a lot different to the larger group, they skew the outcome maybe a bit too much,” she says. “Their genetic information is more rare, and the way the maths compute this inbreeding metric, it gives more power to those rare alleles.”

‘Hybrid vigor’ – or not

In addition, they compared levels of inbreeding with survival in chicks. When the mainland male was added to the Stewart Island population, the expectation was that he would infuse genetic diversity into a new generation of chicks, boosting their health. But that turned out not to be the case: decreased inbreeding did not correspond to improved survival, partly because the less-inbred chicks had unexpectedly high levels of mortality.

“That was probably down to the mainland individuals also being reduced to a small population for a long period of time,” says Foster. The mainland male apparently brought in quite a few detrimental mutations that had been lost from the island population over the years.

The inbreeding study and others like it could help guide conservation strategies moving forward. Kākāpō have a lek mating system, which means that the males all congregate and compete to entice the females. The most popular male will produce the most offspring, and this can tighten the genetic bottleneck even more.

“One male kākāpō had fathered 22 chicks, which is really significant when there’s only 200 left,” says Foster. “Obviously his genetic material spread across the population. He was so successful, we had to translocate him to another island to give some of the other males a chance.”

CITATION

Genomic signatures of inbreeding in a critically endangered parrot, the kākāpō

Yasmin Foster, Ludovic Dutoit, Stefanie Grosser, Nicolas Dussex, Brodie J. Foster, Ken G. Dodds, Rudiger Brauning, Tracey Van Stijn, Fiona Robertson, John C. McEwan, Jeanne M. E. Jacobs, and Bruce C. Robertson

G3 Genes|Genomes|Genetics 2021; jkab307

https://doi.org/10.1093/g3journal/jkab307

A new genome assembly for Antarctic fur seals sheds light on their historic comeback after 19^th century hunting.

In the late 19^th century, the Antarctic fur seal was thought to be effectively extinct. After over a century of overexploitation driven by demand for the seal’s prized pelt, populations at known breeding grounds seemed to have disappeared, making further hunting impossible—and suggesting that the species may even have died out altogether. But in the 1930s, a small breeding population was discovered on South Georgia, a remote island in the southern Atlantic Ocean with no indigenous human inhabitants. Today, the Antarctic fur seal has made a comeback, with a population thought to number as many as two or three million—but a new G3 report by Humble et al. suggests this picture of the seal’s dramatic rebound is incomplete.

As a well-studied species that has undergone a remarkable recovery, the Antarctic fur seal (Arctocephalus gazella) holds great interest for conservation biologists and others seeking to understand the genomic impacts of population changes. In the report, a multinational team of authors describe an improved A. gazella genome assembly and a collection of 677,607 single nucleotide polymorphisms (SNPs), both useful tools for deeper dives into the genetics of the species. Their data also contain clues about how the Antarctic fur seal may have repopulated much of its former range.

Humble et al. found that linkage disequilibrium in A. gazella is on par with that of other vertebrates—a result that may seem strange given that such a severe population bottleneck should increase linkage disequilibrium. However, a separate analysis recently hinted that the population may not have dropped as low as once thought and could have included hundreds of individuals at its minimum. The Antarctic fur seal population also recovered within just a few generations, reducing the amount of time inbreeding and genetic drift would have had to impact linkage disequilibrium.

Although the species has a large, free-ranging population, the researchers found that some individuals were more inbred than others. This may be due in part to the fact that both males and females of the species return to the same breeding grounds each year with great precision—in one study, females were found to return to within one body length of the places they were born. Further, the species is highly polygynous, with one male often siring offspring with over a dozen females in a given season.

Information about fur seal population structure gave the team evidence that A. gazella may have persisted at a small number of the breeding grounds and thus was not limited to South Georgia, where it was first spotted after hunting ceased. Further investigation of how the seal recovered from being critically endangered, including the role of these final holdouts, could provide valuable information to guide conservation of other species facing extinction. And while A. gazella now numbers in the millions, any such insight may one day be important for its preservation, too: climate change and an increase in tourism has begun to put pressure on many Antarctic species, including the resilient fur seal.

CITATION:

RAD Sequencing and a Hybrid Antarctic Fur Seal Genome Assembly Reveal Rapidly Decaying Linkage Disequilibrium, Global Population Structure and Evidence for Inbreeding
Emily Humble, Kanchon K. Dasmahapatra, Alvaro Martinez-Barrio, Inês Gregório, Jaume Forcada, Ann-Christin Polikeit, Simon D. Goldsworthy, Michael E. Goebel, Jörn Kalinowski, Jochen B. W. Wolf, Joseph I. Hoffman
G3: Genes, Genomes, Genetics 2018 8: 2709-2722; https://doi.org/10.1534/g3.118.200171
http://www.g3journal.org/content/8/8/2709

A green, iridescent bee perches on a pink flower, extending its proboscis to reach the sweet nectar inside. He’s not just after a meal—he’s also collecting fragrant substances to store inside his hollow rear legs. Later, he’ll buzz his wings to release the aroma with the hope of attracting a mate. The cover of the September issue of G3 features a photograph of this eye-catching insect: a type of orchid bee called Euglossa dilemma. Orchid bees inhabit the neotropical realm, a region encompassing most of South America, some of Central America, and a tiny fraction of southern North America. There, these bees are some of the most important pollinators of flowering plants.

In the same issue of G3, Brand et al. report a draft assembly of the nuclear and mitochondrial genome of E. dilemma, the first draft genome of any species in the genus Euglossa. The genome revealed several interesting facts about the bees; for example, they have one of the largest genomes of any insect, loaded with repetitive sequences. Their assembly will also be a boon to bee researchers, from those seeking to know more about how to conserve these essential pollinators to those studying bee evolution.

Of particular interest is the evolution of one of many bees’ most fascinating traits: the ability to form intricate social structures. E. dilemma, unlike its close relatives the honeybee, stingless bee, and bumble bee, doesn’t actually live in communal hives. If the male orchid bee succeeds in seducing a female with his foraged scents, she’ll lay their eggs in a small nest of up to twenty cells, where she’ll feed the larvae nectar and pollen. Orchid bees may build their nests near each other, giving the impression of a loosely connected society, but groups of nests don’t form communities like hives. However, daughter bees sometimes stay in their mothers’ nests to help her raise a new generation—a type of social interaction that may have been one of the evolutionary stepping stones toward hives in insects like honeybees. Using the new genome as a starting point, researchers might be able to learn more about the evolution of this complex behavior, increasing our knowledge of many more types of these industrious insects.

CITATION:
Brand, P.; Saleh, N.; Pan, H.; Li, C.; Kapheim, K.; Ramírez, S. The Nuclear and Mitochondrial Genomes of the Facultatively Eusocial Orchid Bee Euglossa dilemma.
G3: Genes|Genomes|Genetics, 7(9), 2891-2898.
DOI: 10.1534/g3.117.043687
http://www.g3journal.org/content/7/9/2891

Towering sugar pine trees dominate the mountain forests of California and Oregon. They are the tallest pine trees in the world, regularly growing to skyscraper heights of over 100 meters. But these forest behemoths are under attack from a very tiny foe: an invasive fungus. White pine blister rust was accidentally introduced to western North America nearly a century ago. Since then, blister rust infections have been threatening the survival and reproduction of sugar pines, harming the ecosystem and industries that depend on them. Conservation efforts have shown that genetic variation contributes to the likelihood that one tree and not another succumbs to infection, but efforts to track down the genes involved have been complicated by the staggeringly huge genome of this giant tree and the arduous tests. The sugar pine genome is ten times the size of the human genome—a whopping 31 billion base pairs. Kristian Stevens and colleagues announced the complete sequence of the sugar pine genome in the December issue of GENETICS, the largest genome fully sequenced to date. Their work, along with a companion paper on the sugar pine transcriptome published in G3, highlights the evolutionary implications of such a massive genome size, as well as revealing candidate genes for blister rust resistance and a promising path to efficient selection of resistant individuals.

Despite its enormous size, the sugar pine genome contains about the same number of protein coding genes as the human genome. No less than 79% of the DNA in the sugar pine genome is made up of transposable elements, which accounts for its enormous size. These genetic parasites are stretches of DNA that exist only to proliferate within a genome. Rather than contributing to the sugar pine’s phenotype, they encode machinery that lets them make copies of themselves at new sites in the genome. Transposable elements are common in all eukaryotic genomes, but in conifers, and especially the sugar pine, they have multiplied to enormous numbers. In the sugar pine genome, the transposable elements are mostly non-functional relics. These genomic leftovers can tell researchers about the evolutionary history of the sugar pine and also provide insights about how genomes size evolves. They also create substantial problems for researchers trying to work with the sugar pine genome.

Transposable elements are highly repetitive, and when they are present in numbers as large as in the sugar pine, they are extremely difficult to sequence. Whole genome sequencing generally works by breaking a genome up into extremely small pieces and then putting them back together one by one. Repetitive genetic sequences make this process incredibly difficult because when the pieces are assembled, all the repeats look the same and end up incorrectly merged into one sequence. To get around this problem, the researchers assembling the sugar pine genome used several strategies. They obtained most of the sequence data from a single haploid pine nut, avoiding the typical complications of sequencing two parental genomes in a diploid individual.They sequenced the transcriptome to identify those sequences that produce proteins, and then used those sequences to assemble the corresponding genes. They also used sequencing libraries specially prepared with the reads known to be large distances away from one another, which is useful in linking larger genomic structures—the big picture. These techniques, along with others, allowed the researchers to build a useful working draft of the massive sugar pine genome.

A twig infected with white pine blister rust. Photo by Marek Argent via Wikimedia.

Sequencing an entire genome, especially one as large as the sugar pine, is an impressive technological achievement. More importantly, however, it is an incredibly powerful research tool in the fight against white pine blister rust. This fungus has been infecting multiple species of white pines in the North America since it was first introduced from Asia around the turn of the century. White pine blister rust is a slow killer, taking years to destroy a large tree. An infection begins when fungal spores land on the surface of the tree and begin to germinate. They grow through openings into the twigs and branches, and very slowly make their way towards the main trunk of the tree. The infected branches swell up and large sacks of rusty orange-red spores burst through the branches. The fungal infection causes cankers, which prevents the tree from sending water and nutrients to its damaged limbs. Eventually, these limbs will die. If cankers form on the main trunk, the entire tree may die.

Researchers and forest managers have been looking for a way to fight the spread of white pine blister rust for a long time. Some rare sugar pines carry genetic resistance to white pine blister rust, and have been used in reforestation efforts. In the 1970s, these rare individuals were used to identify a major locus of resistance called Cr1, but the daunting size of the sugar pine genome made further analysis difficult. Using this new genome sequence, Stevens and colleagues were able to make a breakthrough in identifying this gene. They used the small amount of genetic information already known to find large Cr1-associated segments and identify previously unknown SNPs that are closely associated with resistance. These markers are a powerful tool that can be used to quickly and cheapy identify trees that carry the resistant allele without waiting for the results of slow and expensive infection assays. Resistant trees can then be harvested for seeds to be used in reforestation. Now armed with a roadmap, scientists can search the sugar pine genome for the secrets that may help save these iconic trees and the ecosystems that depend on them.

Stevens, K. A., Wegrzyn, J. L., Zimin, A., Puiu, D., Crepeau, M., Cardeno, C., Paul, R., Gonzalez, D., Koriabine, M., Holtz-Morris., A. E., Martínez-García, P. J., Sezen, U.U., Marçais, G., Jermstad, K., McGuire, P. E., Loopstra, C. A., Davis, J. M., Eckert, A., deJong, P., Salzberg, S. L., Neale, & Langley, C. H. (2016). Sequence of the Sugar Pine Megagenome. Genetics, 204(4), 1613-1626. DOI: 10.1534/genetics.116.193227

http://www.genetics.org/content/204/4/1613.abstract

Gonzalez-Ibeas, D., Martinez-Garcia, P. J., Famula, R. A., Delfino-Mix, A., Stevens, K. A., Loopstra, C. A., Langley, C. H., Neale, D. B., & Wegrzyn, J. L. (2016). Assessing the gene content of the megagenome: sugar pine (Pinus lambertiana). G3: Genes| Genomes| Genetics, 6(12), 3787-3802. DOI: 10.1534/g3.116.032805

http://www.g3journal.org/content/6/12/3787.short

The sex of many reptile species is set by temperature. New research reported in the journal GENETICS identifies the first gene associated with temperature-dependent sex determination in any reptile. Variation at this gene in snapping turtles contributes to geographic differences in the way sex ratio is influenced by temperature. Understanding the genetics of sex determination could help predict how reptiles will evolve in response to climate change.

In crocodiles, alligators, and certain lizard and turtle species, an embryo can become either a male or a female depending on the temperatures it experiences while in the egg. Rapid climate change may threaten the future of some of these species by skewing the sex ratio. For example, by some estimates temperature rises over the next century will cause painted turtles to produce only females. Such species may also evolve in response to climate change. Biologists are trying to understand how these animals will be affected by and adapt to rising global temperatures.

But little is known about how this temperature-dependent switch between ovaries and testes is regulated. To look for clues to the molecular mechanisms behind this process, study leader Turk Rhen (University of North Dakota) and his colleagues investigate how genes influence sex determination in common snapping turtles. The advantage of focusing on this rugged-looking North American native is that sex is determined in a brief five-day window during the 65-day egg incubation: the temperature-sensitive period. If the incubation temperature during the temperature-sensitive period is changed from a “male-producing temperature” (26.5°C or 79.7°F) to a “female-producing temperature” (31°C or 87.8°F), all the eggs will hatch into females.

Snapping turtle adult. Photo: Turk Rhen

In previous work, the team identified a gene—CIRBP—that is activated within 24 hours of such a temperature shift. Two days later, several genes known to be involved in ovary or testes development are either activated or repressed. The new study confirmed that CIRBP is expressed at the right time (very early in the temperature-sensitive period) and the right place (the gonads) to be involved in specifying sex.

To test whether this hypothesis is correct, the researchers investigated DNA sequence variation at the CIRBP gene, and whether it influenced the chances of an individual turtle becoming male or female.

They found that some of the turtles carried a slightly different version of CIRBP: at a specific position in the sequence, an “A” in the four-base DNA code was substituted with a “C”.

This change rendered the gene unresponsive to temperature: instead of being induced by the female-producing temperature, the “C” version of the gene remained at steady levels.

Individuals carrying this unresponsive “C” version were more likely than average to be male. This single-letter DNA difference between turtles could explain around a quarter of the genetic variability in sex determination temperatures, which suggests that the activation of CIRBP is a crucial event in specifying sex.

Remarkably, this CIRBP variant may partly explain a curious fact about snapping turtles: the sex ratio in populations from different latitudes responds differently to temperature. For example, if you collect eggs from snapping turtles in Minnesota and Louisiana and incubate them all at 27°C (80.6°F) in the lab, the eggs collected in the North will produce nearly all males, while those from the South will produce mostly females. This variation suggests subpopulations of the species have evolved and adapted to their local climate.

The team found that the “C” version of CIRBP was more common in turtles from northern Minnesota than those from 250 miles away in southern Minnesota, and it was not detected in a population from even further south, in Texas. Though this is only a small sample of locations, the trend is consistent with the sex determination pattern in each population: the “A” version (which makes turtles more likely to be female) was more common in populations that produce females at a lower temperature.

The protein encoded by the CIRBP gene (cold-inducible RNA-binding protein) is likely involved in sensing temperature and converting it into a developmental signal to trigger the formation of either testes or ovaries, says Rhen. Studies from other organisms suggest that this protein can regulate temperature-dependent processes by binding to the RNA “messages” produced by specific genes.

“CIRBP seems to play a crucial role in sex determination,” says Rhen. “The striking part is that we see a consistent association across multiple levels of biology: the variation at the DNA level influences the gene’s activation (expression into RNA messages), which is in turn correlated with whether an individual turtle becomes male or female. That association with sex holds whether we look at individuals or families, and we even see differences at the population level.”

But CIRBP is not the only gene important for specifying sex in snapping turtles, the data show. “There is a common misconception that there must be a single “magic bullet” gene that determines sex in response to temperature,” says Rhen. “Our data suggests that multiple temperature sensors control sex by acting together. We’re trying to identify the other components of this system and to determine how they interact to influence sex. Better understanding variation at these genes may one day allow us to predict how reptile species will evolve under a new climate regime.”

CITATION

A Novel Candidate Gene for Temperature-Dependent Sex Determination in the Common Snapping Turtle

Anthony L. Schroeder, Kelsey J. Metzger, Alexandra Miller, Turk Rhen

GENETICS May 1, 2016 vol. 203 no. 1 557-571; DOI:10.1534/genetics.115.182840

http://www.genetics.org/content/203/1/557

In 1996, when I started researching the conservation genetics of New Zealand’s critically endangered parrot, the kākāpō (Strigops habroptilus), little was known of the species’ genome.  On many occasions after a long day in the molecular lab on the hunt for an elusive gene, I found myself imagining that I had the complete genome of kākāpō at my fingertips. Fast forward 20 years, and that dream has been realised with the sequencing of the kākāpō genome by researchers at Duke University and PacBio, as part of the Bird 10,000 genomes project. It was like Christmas the day the whole genome landed in my inbox (22 January 2016). Elusive kākāpō genes could no longer hide from me and the possibilities for kākāpō conservation and species recovery seemed endless.

Many people will be familiar with the species thanks to a kākāpō called Sirocco (the species’ “spokesbird,” who tweets @Spokesbird) and his unusual mating habits. Stephen Fry’s narration of Sirocco mating with Mark Carwardine’s head on YouTube has had over 6.7 million views. For many others like me, Douglas Adams introduced kākāpō to them in the moving account of the species’ plight in the book Last Chance to See. Historically a widespread species, the kākāpō reached a low of only 51 birds in 1995, and the world population now stands at a hard-won 125 individuals. To maintain genetic diversity and ensure species recovery, all birds are now intensively managed by a team of dedicated conservationists.

Photo courtesy Andrew Digby / DOC

Conservation genomics – the application of genomic analysis to the management of threatened species – is a fast-emerging field propelled forward by the dramatic advances in genome sequencing.  These technological developments have also led to considerable cost savings, and we are to the point where we can once again dream big for kākāpō. In a world first, the Kākāpō 125 Genomes Project is sequencing the genome of every surviving individual of a species.  

This ambitious project is a collaboration among researchers from the New Zealand Department of Conservation’s Kākāpō Recovery Group, the University of Otago (New Zealand), Duke University (USA), the Genetic Rescue Foundation and NZ Genomics Ltd.

For the Kākāpō 125 Genomes Project, we’re using the high-quality reference kakapo genome developed by Duke University and PacBio as a scaffolding to assemble all other 124 kakapo genomes against. This project is being funded by the Genetic Rescue Foundation, a not-for-profit organization dedicated to advancing the scientific techniques required to prevent species extinction through genetic intervention. The Genetic Rescue Foundation makes use of the latest crowdfunding and online scientific services such as Science Exchange and Experiment.com to fund and conduct research. The Genetic Rescue Foundation’s latest crowdfunding campaign is currently in progress and in just 20 days has raised over $32,000 toward a target of $45,000.

A genomics approach to kakapo conservation will be a great boost to the recovery program. A genome-wide understanding of genetic variation will help to develop breeding strategies to retain variation at genes important for species persistence, such as the immunity genes and their role in kākāpō diseases. We will also be able to explore the genetic basis of infertility in kakapo; only 60% of eggs hatch (normally this should be about 90% in birds) and sperm abnormalities contribute to infertility. Solving the issue of infertility will greatly aid species recovery by maximising reproductive effort.

Beyond being of considerable significance to kākāpō conservation, the Kākāpō 125 Project will make an important contribution to the emerging field of Conservation Genomics. Our project will provide a genome dataset for every member of an entire species, serving as a much needed case study to further develop analytical pipelines and methodologies. Only with studies such as ours will Conservation Genomics’ true potential be realised.

Bruce Robertson is an Associate Professor studying the conservation genomics, molecular ecology and wildlife management of New Zealand native species in the Department of Zoology at the University of Otago, Dunedin, New Zealand.

The views expressed in guest posts are those of the author and are not necessarily endorsed by the Genetics Society of America.

Amphibians around the world have been devastated by the spread of the deadly fungus Batrachochytrium dendrobatidis (Bd). But although many populations have been decimated, others have survived the same threat. One reason for such different outcomes is variation in virulence between Bd isolates. In the latest issue of G3, Refsnider and Poorten et al. investigate Bd virulence changes by taking advantage of the rapid evolution that can take place in the lab.

The authors studied a strain isolated in 2005 from an infected frog found in the El Yunque rainforest, Puerto Rico; one isolate was frozen soon after, while a second isolate was maintained in laboratory culture for six years. Today, the isolate that has grown used to lab life is significantly less lethal to frogs. Studying the genetic basis of this shift may shed light on how virulence is attenuated in the wild, particularly because many of the confounding factors inherent in natural systems can be controlled for by comparing two isolates from a single lineage.

The authors resequenced the pair of isolates to look for genomic changes underlying the phenotype difference. They found that the two strains had diverged at a rate of 1.6 x 10^-5 mutations per site per year, which is faster than most fungi but similar to other fungal pathogens. The less virulent isolate had lower chromosomal copy numbers than its wilder cousin, suggesting one way that virulence can be rapidly lost. The extra chromosome copies needed for virulence may become costly to the pathogen once normal selection pressures have been relaxed.

The speed of Bd’s change in the lab suggests this pathogen is capable of rapid adaptation to new conditions, consistent with its alarming spread across habitats and continents.

CITATION

Refsnider, J. M., Poorten, T. J., Langhammer, P. F., Burrowes, P. A., & Rosenblum, E. B. (2015). Genomic Correlates of Virulence Attenuation in the Deadly Amphibian Chytrid Fungus, Batrachochytrium dendrobatidis. G3: Genes| Genomes| Genetics, 5(11), 2291-2298 doi:10.1534/g3.115.021808

http://www.g3journal.org/content/5/11/2291.full

In 2009, after five years parching under the arid blue skies of Calcena in northeastern Spain, dozens of neat rows of maritime pine seedlings had grown unevenly. Some of the seedlings had died years ago, some were stunted but hanging on, while others grew tall and green.

The trees were not in their native soil. They had been grown from seeds collected at 19 sites around Spain, Portugal, France, and Morocco, and their growth was being monitored at a single site with an extreme climate to help predict the future of their species.

The experiment was designed to improve models that forecast where forests will grow as the southern European climate grows hotter and drier, and promises to help forestry managers choose tree stocks and decide where to focus reforestation efforts. In the March issue of GENETICS, Jaramillo-Correa et al. reported the results, identifying a handful of SNPs that can be used as predictors of maritime pine survival under different climatic conditions.

The maritime pine (Pinus pinaster Aiton) grows widely in southwestern Europe and parts of northern Africa. But the tree’s important economic value and ecological roles in the region may be at risk as the changing climate threatens the more vulnerable forests and the productivity of commercial plantations.

The species range is expected to move northward as the climate changes, says study leader Santiago González-Martínez, from the Forest Research Centre of Spain’s Institute for Agricultural Research (CIFOR-INIA). “But many populations in the Mediterranean region are already at risk, and we don’t necessarily know what genetic resources and adaptations we will lose if they disappear,” he says. “Another big problem for commercial plantations is that their tree stocks have been intensively bred for productivity, but not for resistance to drought. Forestry breeding cooperatives are very interested in introducing trees that are more resilient to climate change, with increased genetic diversity.”

Maritime pine common garden test site in Calcena, Spain. Image credit: Eduardo Notivol

Range-shift models are key tools for managing forests as the climate changes. These forecasts are based mainly on ecological and physiological data, however, and don’t take into account two major influences on a forest’s fate: genetics and evolution. Genetic differences between tree populations mean that forests vary in the degree to which they cope with changing conditions. Natural selection will also influence the prevalence of such genetic variants as the climate shifts.

Genetic effects can drastically change range-shift predictions, says González-Martínez. The maritime pine project sought to identify and quantify such effects in a way that could be readily incorporated into existing models.

To track down genetic variants that affect maritime pine fitness in different climate conditions, the team decided on a candidate gene approach, which is considerably faster and cheaper than surveying the large and complex maritime pine genome. Pine researchers from around the world pooled their expertise to yield a list of more than 300 SNPs in 200 candidate genes. “It was really a community effort, drawing on 15 years of research across many labs,” said González-Martínez.

From this candidate list, the team tested whether any of the SNPs were associated with climate variables across 36 natural populations, after correcting for geographic patterns in SNP frequency due to the different demographic histories at each site. Eighteen of the candidate SNPs showed significant associations with climate factors. These variants affected many different biological processes, including growth and response to heat stress.

The researchers then looked for evidence that these SNPs are important for fitness. They planted over 6,000 seedlings from 520 families and 19 locations together in the “common garden” in Calcena, where the climate falls at the extreme dry end of the species’ range. After five years, tree survival was significantly correlated with the frequency of SNP alleles predicted to be beneficial in the Calcena climate. In other words, the seedlings that were still thriving after five years in their new home tended to be the ones genetically well equipped to survive the harsh climate.

These results demonstrate the feasibility of this relatively fast approach of finding and confirming genetic variants associated with climate. Now that they have shown the method works, González-Martínez and his colleagues are expanding the project to cover more genes and more traits. “The single biggest climate change threat to pine forests is the increased frequency of wildfires, so we’re searching for variants that affect fire tolerance,” he says. They are also planting common gardens at many different locations—growing thousands more little seedlings whose fate will help geneticists predict the maritime pine’s future.

Read the press release: http://www.genetics-gsa.org/media/releases/GSA_PR_201503_pine.html

CITATION:

Jaramillo-Correa J.P., D. Grivet, C. Lepoittevin, F. Sebastiani, M. Heuertz, P. H. Garnier-Gere, R. Alia, C. Plomion, G. G. Vendramin & S. C. Gonzalez-Martinez & (2014). Molecular Proxies for Climate Maladaptation in a Long-Lived Tree (Pinus pinaster Aiton, Pinaceae), Genetics, 199 (3) 793-807. DOI: http://dx.doi.org/10.1534/genetics.114.173252 
http://www.genetics.org/content/199/3/793.full