Researchers generate high-quality genome assembly for taro

It is the first genome assembly of taro that has been associated with genes that underpin resistance to taro leaf blight, information that may enhance taro breeding programs.

By Susan Enright

Man in taro field.
Genetic researcher Roshan Paudel in taro field. Courtesy photo.

For decades, a team of researchers from the University of Hawai‘i at Hilo and UH Mānoa crossbred taro species that show resistance to taro leaf blight (TLB) in the hopes of introducing disease resistance traits through breeding programs. All Hawaiian varieties of taro are vulnerable to TLB, a highly infectious plant disease caused by the oomycete pathogen Phytophthora colocasiae.

But although a Palauan taro resistant to TLB crossbred with Hawaiian taro produced some very good hybrid varieties that are currently used in commercial poi, other trials resulted in varieties unacceptable for commercial poi processors.

Now the researchers have turned their sights on genomics, the painstaking study of nucleotide sequences in the chromosomes of an organism. The paper on this genomic work, “Taro Genome Assembly and Linkage Map Reveal QTLs for Resistance to Taro Leaf Blight,” was published online in the journal G3: Genes, Genomes, Genetics on Aug. 1, 2020.

The project was done to develop more tools for managing taro leaf blight. In the study, the researchers generated a high-quality genome assembly for taro, a root crop that is widely cultivated in tropical regions and is important for food security. The results are intended to inform studies of the origin, evolutionary history, and breeding of the South Pacific crop. It is hoped the genome may also stimulate new genetic insights into this important tropical species, especially in regard to taro leaf blight.

M. Renee Bellinger in lab
M. Renee Bellinger

“Overall, this project was a large team effort, involving three universities and U.S. Department of Agriculture researchers,” explains M. Renee Bellinger, a post-doctoral affiliate faculty and adjunct assistant professor of bioinformatics and genomics at UH Hilo who conducted genomic research on the project.

In addition to Bellinger, also on the genomic research team from UH Hilo are Michael Shintaku, Martin Helmkampf, Steven Starnes, and Lucas Kambic; from UH Mānoa are Susan Miyasaka, Roshan Paudel, Michael B. Kantar, and Thomas Wolfgruber; from University of Tennessee is Kurt Lamour; and from the U. S. Department of Agriculture are Scott Geib and Sheina Sim.

“This project was able to combine a huge amount of information, it brought together large scale sequencing with linkage mapping to create a genome that is easy to use and provides biological insight,” says Kanter, an assistant professor at UH Mānoa specializing in plant breeding and genetics.

The long fight against taro leaf blight

Taro, or kalo in Hawaiian, is considered a sacred plant in Hawai‘i and has been a part of Hawaiian agriculture and society for centuries, and is believed to have been brought by the first Polynesians who migrated to Hawai‘i. Somewhere between 150 to 300 varieties were once present on the Hawaiian Islands, but many have been lost, with about 70 varieties remaining.

The recently published work was conducted as part of the long-term research project led by UH Hilo Professor of Plant Pathology Shintaku and UH Mānoa Professor of Agronomy Miyasaka.

Taro lef with edges brown and falling away.
Taro plant with taro leaf blight. Courtesy photo, click to enlarge.

“We started this project to develop tools for managing taro leaf blight,” says Shintaku. “TLB has been causing serious problems in Hawaiian kalo fields since the 1920s. Samoa didn’t have TLB until about 1990, and then suffered dramatic losses.”

Shintaku explains Phytophthora colocasiae, the pathogen that causes TLB, is a fungus-like organism closely related to Phytophthora infestans. “Interestingly,” he says, “P. infestans causes late blight of potato and triggered the Irish potato famine of 1845, and a mass migration of Irish to the Americas.”

UH researchers have been trying to develop resistance to taro leaf blight for decades, and field trials with imported Palauan varieties showed promising results. The Palauan varieties had less TLB damage and subsequently grew faster and larger. However, the poi produced from these varieties was unacceptable for commercial poi processors.

A breeding program was started, to incorporate Palauan resistance to TLB into Hawaiian taro. This effort produced some very good hybrid varieties that are currently used in commercial poi.

“Recent breeding efforts at the Waiakea Experiment Station were conducted by Christopher “Popo” Bernabe, and we used many hundreds of the plants he produced in our work,” explains Shintaku.

The researchers also worked with a local kalo farmer so knowledgeable in the crop that he was consulted by lawmakers, educators, botanists, researchers, and Native Hawaiians who needed help identifying, cultivating, or protecting the sacred plant.

“Uncle Jerry Konanui was the president of Hui Kalo Moku o Keawe, and he worried that distribution of hybrid kalo would confound the efforts of those, like Hui Kalo Moku o Keawe, trying to preserve the ancestral Hawaiian varieties,” Shintaku says. “He encouraged us to develop DNA fingerprinting for the Hawaiian varieties, and helped us greatly with collecting and identifying the varieties. We were pleased to share our preliminary results with him in 2014. Sadly, we lost Uncle Jerry in 2017.”

Uncle Jerry and Mike Shintaku in cafe with laptop open on table.
From left, Jerry Konanui and Mike Shintaku, when Prof. Shintaku shared preliminary DNA fingerprinting with Uncle Jerry on March 6, 2014. Courtesy photo from Mike Shintaku.

The genomic research

While Shintaku and Miyasaka have worked toward this type of genomic project for decades, only recently have the cutting edge genetic tools become accessible to conduct genomic research with this level of depth.

Shintaku explains the science.

Michael Shintaku
Michael Shintaku

“Single-nucleotide polymorphisms or SNPs are genetic markers and the recently published work was a SNP-discovery effort,” he says. “If you look at a 100-nucleotide segment from one individual and compare it with the corresponding segment from another individual you might find no difference,” explains Shintaku. “If, however, you find one nucleotide that is different, you have a SNP. SNPs are useful for DNA fingerprinting, and we used them for differentiating taro varieties. However, if traits like disease resistance can be linked to particular SNPs they can be used to greatly accelerate a breeding program.”

Research team member Roshan Paudel, who recently completed his master of science degree at UH Mānoa under the mentorship of now colleague Michael Kantar, is now a doctoral student at UHM. Paudel looked for linkage between SNPs and plants with TLB resistance. He looked at hundreds of SNPs, searching for ones linked to quantitative trait loci (QTLs), which can additively contribute to resistance.

“The DNA sequence of the taro genome would tell us the location of the SNPs and tell us which ones are most important, shedding light on some important characteristics of kalo,” explains Shintaku. “A genome sequence is also a valuable global resource for taro research.”

As part of Renee Bellinger’s post-doctoral fellowship in bioinformatics and genomics at UH Hilo, she worked with Shintaku (based at UH Hilo College of Agriculture, Forestry and Natural Resource Management), Miyasaka and Kantar (both from UH Mānoa Department of Tropical Plant and Soil Sciences), to sequence the taro genome and analyze genome characteristics.

Paudel applied results from a next-generation sequencing data set to identify regions of the genome associated with taro disease resistance to the pathogen that causes taro leaf blight.

Bellinger identified regions of the genome with disease resistance genes, and matched those to Paudel’s results. Some of the genomic regions associated with disease resistance were in close proximity to the disease resistance genes.

Meanwhile, team member Lucas Kambic completed his bachelor of science degree at UH Hilo, after which he was hired by Shintaku to do the genome sequencing using a plug-in genome sequencer kit made by Oxford Nanopore. The kit can be used in any lab, and the sequencer plugs into a typical desktop computer. This is one type of emerging technology that places genome sequencing within reach of any laboratory.

Starnes worked with Kambic and performed the Nanopore genome assembly.

Bellinger explains that Miyasaka’s group screened more than 2,500 plants, with one cross that showed particularly promising results. Paudel focused on that TLB resistant cross for his master’s thesis. Paudel matched next-generation sequencing data to the disease resistance trait data and identified 520 genetic markers. Next, he tested those markers for association with TLB disease resistance, and identified 10 that associated with that trait. He also used these markers to construct a linkage map, which identifies SNP markers likely to originate from the same chromosome.

Another aspect of this project is to shed light on the genetic underpinnings of plant disease resistance by sequencing the genome of one of the Hawaiian varieties, Moi, a grandparent to the disease resistant cross studied by Paudel. The genome analysis, led by Bellinger, revealed that plant disease resistance genes occur in clusters across the genome, and that two of the genetic markers associated with TLB resistance occur in the proximity of plant disease resistance genes.

“This is the first genome assembly of taro that has been associated with genes that underpin resistance to taro leaf blight, information that may enhance taro breeding programs,” Bellinger says.

She continues, “To me, this project exemplifies synergies that can be achieved by research teams working collaboratively towards a common goal. I was able to use Roshan’s linkage map to improve the genome assembly, and Roshan was able to use the genome assembly to place his genetic marker findings in a broader context.”

Shintaku says the team was also very fortunate that Vernon Oi, a lead scientist in monoclonal antibodies, DNA testing and modification who is the second highest royalty producer at Stanford, was also interested in developing this resource. “His support made possible the sequencing effort,” says Shintaku.

“It is an exciting time to be working in the field of genomics,” says Bellinger. “Roshan and I share the sentiment that it is a privilege to work with such a culturally and historically important crop.”

 

Story by Susan Enright, a public information specialist for the Office of the Chancellor and editor of UH Hilo Stories. She received her bachelor of arts in English and certificate in women’s studies from UH Hilo.

 

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Michael Shintaku, Professor of Plant Pathology