Group of researchers with Brazilians sequence the Arabica coffee genome

O coffee is one of the most traded commodities in the world and the Coffea species arabica It is the most consumed among the approximately 130 that exist. It is the result of the fusion of two other species: Coffea canephora (known in Brazil as conilon coffee or Robusta) and Coffea eugenioides. Over the past ten years, almost every major commodity in the world has had a reference genome sequenced, but coffee has only recently joined this list.

The reference genome is crucial for the development of cultivars that are more adapted to climate change and resistant to diseases. By sequencing the reference genome of Arabica coffee in an unprecedented work, a consortium of scientists managed to select genes possibly responsible (candidate genes) for coffee's resistance to rust and other diseases. In parallel, it identified the expression of some genes related to the aroma of arabica.

“With knowledge of the genome it is possible to obtain information that allows us to go both ways: the development of varieties through crossbreeding, that is, as a reference to guide us in future crosses that produce new varieties; and more direct interventions, such as modifying a gene specifically”, summarizes Douglas Domingues, currently a researcher at the Plant Genomics and Transcriptomics Group at the Luiz de Queiroz College of Agriculture at the University of São Paulo (Esalq-USP) and one of the authors work (developed when I was still working at the Universidade Estadual Paulista, Rio Claro campus).

According to him, there was a certain race to sequence this genome. “The price for sequencing fell a lot and coffee was one of the few commodities that did not yet have the reference genome sequenced. There were other groups trying, and there was work published shortly before ours. But most used the standard strategy: they chose an interesting plant to grow and sequenced its genome,” he reports.

The group that Domingues is part of has sequenced a plant that, from an agronomic point of view, is of no interest, but from a genetic point of view has a lot to offer. “The advantage of our reference genome is that it is derived from a 'dihaploid' individual [tem dois conjuntos de cromossomos]. This leads to a homogeneous reference genome, which will be a superior standard for future research”, explains Patrick Descombes, coordinator of the work and senior expert in genomics at the Nestlé Institute of Food Safety & Analytical Sciences. He explains that Arabica coffee is a tetraploid (four sets of chromosomes): it has two genomes within one, because it is the fusion of two other species.

By sequencing a dihaploid derived from Arabica coffee, compared to a common tetraploid cultivar, scientists get a clearer and more simplified view of the genome. This allows variations between similar genes to be identified with greater precision, facilitating the use of molecular information for breeding studies.

In this work, the group was able to determine more accurately the time in which this fusion took place: at most, 600 thousand years ago, C. canephora and C. eugenioides merged into this tetraploid hybrid, which followed its evolutionary path. “We reached this conclusion using DNA information from Arabica, Robusta and Eugenioides: we were able to make a more accurate inference, as, previously, this interval was dated between 50 thousand and 1 million years. We reduced this window to 350 to 600 thousand years”, reports Domingues.

The article, published in Nature Genetics this Monday (15/04), was the result of a consortium of scientists from more than ten countries, including Brazil, which participated with more than one institution. In the case of Domingues, participation was partially financed by FAPESP through a Young Researcher project and a Post-Doctoral Fellowship granted to Suzana Tiemi Ivamoto-Suzuki, also author of the article.

“We used the reference sequence to understand the diversity that exists in wild Arabica coffees, from the African region of origin, and compare this with the Arabica coffees that are cultivated today”, says the scientist from Esalq-USP, explaining that the group did a resequencing of Arabica coffee varieties planted in different parts of the world, and also of wild specimens collected in the forests of Ethiopia, and managed to understand the difference between wild and cultivated ones.

To gain a genomic perspective on the evolutionary history of Arabica, the consortium sequenced 46 accessions, including three Robusta, two eugenioids and 41 Arabica. The latter included an 18th century type specimen (the physical specimen that the author of the taxon designated at the time of description as being the material on which it was based), 12 cultivars with different breeding histories, the Timor hybrid (a spontaneous cross of arabica with the pest-resistant robust variety of C. canephora) and five of its backcrosses with arabica and 17 wild plus three wild/cultivated accessions collected from the eastern and western sides of the Great Rift Valley in Ethiopia.

“We use the latest genomic technologies, that is, long reads from the PacBio system [para sequenciamento de genes] high-fidelity and proximity binding with Illumina short reads [um sistema integrado para análise de variação genética e função biológica] to generate chromosome assembly. This combination led to an assembly at the chromosomal level of very high quality and integrity”, highlights Descombes.

Illnesses

According to the Esalq-USP professor, among the cultivated species, something very important for improvement was the introduction of genes for resistance to coffee leaf rust. “In the 1930s, Brazil played an important role in this regard. And the IAC [Instituto Agronômico de Campinas] is a pioneering center for studies and improvement. IAC researchers provided us with plants that predated the institution's breeding program, which dates back to the 1930s. Disease-oriented breeding emerged between the 1960s and 1970s, with the main work being to cross a rust-resistant Arabica plant, the called Timor hybrid, with plants cultivated in several countries so that the new varieties were resistant. But it was not known which genes were responsible for resistance.”

The Timor hybrid was discovered in the fields of Timor Island in the 1920s and is naturally resistant to rust and other diseases. “In addition to rust, coffee berry disease, coffee berry borer and coffee stem borer are three other important pests that affect production in many regions of the world. Climate change is also a key concern in controlling pests and diseases, as they allow them to spread to new regions. The trade of green coffee beans between different regions is another factor that can facilitate the spread of certain pests and diseases to new areas”, reveals Maud Lepelley, manager of the Plant Genetics and Chemistry group at the Nestlé Institute of Agricultural Sciences.

In the work published now, the group managed to find sets of genes that, in the literature, were already related to disease resistance and that are only present in post-breeding varieties. “Somehow, the Timor hybrid managed to receive these resistance genes and now we know which ones. There are dozens, but we have reduced this search space. Arabica coffee has 69 thousand genes; we're down to just under 30 genes. Being able to identify these candidate resistance genes, previously unknown, is an unprecedented feat in our research”, reveals Domingues.

But the work is still far from over, as these genes have to be tested. “More research will be needed to identify and create varieties resistant to these and other coffee pests and diseases”, says Lepelley.

The consortium also managed, using molecular genetics, to make a triple separation: showing that the genetic diversity of wild plants in Ethiopia is distinct from that of coffee cultivated today, probably due to a bottleneck and domestication effect, as few plants were selected for this purpose. process. “We show here that genetic diversity was already very low among wild specimens due to multiple pre-domestication bottlenecks and that the genotypes selected for cultivation by man, both the ancient local Ethiopian cultivars and the more recent ones, were already somewhat mixed between lineages. divergent”, say the scientists.

Aroma

In parallel, the Domingues group managed to observe some occurrences related to the expression of genes linked to the quality of coffee, especially aroma. They studied terpene synthase enzymes, which in plants are related to defense against insects, in addition to a gene linked to lipid compounds in coffee, which encodes fatty acid desaturase 2.

“We observed, in an Asian variety of Arabica, that the genes linked to aroma and flavor are more expressed in the fruits by the C. eugenioides subgenome than by the other progenitor. In other words: one of the genomes contributes more than the other to the sensory characteristics of the drink. What we ask ourselves now is: does this apply to all the varieties we sequence, both pre and post-breeding?”, says Domingues.

“This study clarifies how interactions between C. canephora and C. eugenoides genes are associated with characteristics of Arabica coffee, such as aroma. Elucidating gene interactions helps improve our knowledge of the genetic mechanisms underlying important Arabica traits, a fundamental prerequisite for developing new varieties that will ensure the production of coffee beans for future coffee products,” says Lepelley.

A spin-off of the work is already underway, according to Domingues. “I have just started another project, which is an offshoot of this first effort in partnership with French researchers who were part of this consortium. Let's now analyze the uncultivated species of coffee. We want to know the genome of non-cultivated coffee species, but which contain relevant characteristics in a climate change scenario. We are focusing on sequencing species that are more climate resilient. Knowing what genes they have that Arabica coffee doesn't have and makes them resistant to the climate. Eventually, we could introduce or modify genes through gene editing to make crop species more resistant.”

The article The genome and population genomics of allopolyploid Coffea arabica reveal the diversification history of modern coffee cultivars can be accessed at: www.nature.com/articles/s41588-024-01695-w.