Science uses gene editing to develop blast-resistant rice

Gene editing via the CRISPR/Cas9 technique has been used to give blast resistance to rice cultivar BRSMG Curinga in a study by Embrapa Genetic Resources and Biotechnology. Once launched in 2005, BRSMG Curinga was replaced in the market after becoming susceptible to the disease —which is caused by the fungus Magnaporthe oryzae— over the years. The technology resulting from knocking out two target genes related to blast is still in its testing stage, but has the potential to reach the market in a few years.

Photo: Cláudio Bezerra (Researcher Angela Mehta at a greenhouse)

According to the researcher Angela Mehta, once cultivars like BRSMG Curinga are released to the market as resistant or tolerant to blast, they become susceptible in a few years due to the fact that the fungus’ variability is very high. This is a problem for both rainfed and irrigated rice cultivation. The study compared a susceptible rice genotype with a resistant genotype in search of proteins that become more abundant in the susceptible plant after the pathogen infects it.

″We identified a group of proteins that is potentially involved in such susceptibility, and selected corresponding genes for knockout by CRISPR,″ Mehta states. The term ″knockout″ means removing the function of a given gene, which in turn stops producing the functional protein. For the researcher, the study also shows the importance of prospecting genes of agronomic interest using omics techniques (a set of molecular tools that help in understanding the different biological molecules that give functionalities to an organism, such as genomics, transcriptomics, proteomics and metabolomics).

They identified three genes that are capable of such knockout, two of which were validated during Fabiano Távora's sandwich doctorate, which was supervised by Mehta and carried out at the French Agricultural Research Centre for International Development (Cirad).

Knockout

Photo by: Cláudio Bezerra (researcher Angela Mehta)

By knocking out those genes in the Kitaake rice variety, which is considered a model, Távora found that the plant became slightly more resistant to blast in comparison with the unedited variety. ″When he returned from France, we used one of those genes as a target and selected two others to create a combination of targets to knock out two genes at once, in two different builds in the cultivar BRSMG Curinga″, Mehta explains. The study resulted in two blast-resistant rice lines (one for each build) presenting mutations for the two target genes.

According to the researcher, the knockouts of two genes were held at the same time to enhance resistance and obtain a better response. She highlights that the results so far have been obtained in a greenhouse, where the lineages were challenged with an isolate of the fungus. ″The next step is to challenge them with other isolates to see if this resistance will be maintained″, she adds.

Mehta also notes that BRSMG Curinga was chosen because it is a transformable genotype and because it presents agronomic characteristics closer to the producer's interest when compared to model cultivars, such as Nipponbare and Kitaake.  ″These varieties do not have the level of improvement for highlands of Curinga, which was adapted to the conditions of the Brazilian Cerrado″, he points out. 

For the researcher, the research findings offer an important source of genetic resistance to blast, unlike what we have today, to be incorporated into elite lines, varieties and even into lineages of populations for recurrent selection in Embrapa’s rice breeding program.

The study has been conducted in partnership with researchers Raquel Mello and Adriano Castro, from Embrapa Rice and Beans, in a project led by Angela Mehta, funded by Embrapa with the support of an Embrapa-Monsanto project led by the researcher Márcia Chaves, from Embrapa Temperate Agriculture.

Blast

Photo by: Raquel Neves (BRSMG Curinga non-edited rice plants showing typical blast symptoms)

Blast, which is caused by the fungus Magnaporthe oryzae, is considered the most destructive rice disease and occurs throughout Brazil. As losses are variable, they are higher in rainfed rice fields in Midwestern Brazil, and can compromise up to 100% of the yield in years of epidemic attacks. It also attacks several grasses that are common in rice and wheat fields.

The main sources of primary contamination are infected seeds and crop remains. Secondary infection originates from sporulation lesions on infected leaves. All the stages of the disease cycle are highly influenced by climate factors. In general, it requires high temperatures (from 25 °C to 28 °C) and humidity above 90%.

Symptoms on the leaves start with the formation of small brown necrotic lesions, which grow in size and become elliptical, with brown margins and a gray or whitish center. Under favorable conditions, the lesions cause death of the leaves and, often, the entire plant.

The characteristic symptoms in the plant’s knots are brown lesions that can affect the regions of the stem that are close to the attacked knots. A node infection at the base of the panicle (rice plant inflorescence) is known as collar blast, as a brown lesion surrounds the nodal region and causes plant strangulation.

The plant may present stunting (an anomaly that occurs when the grains do not develop correctly). The panicles turn whitish and are easily identified in the field. Many parts of the panicle, such as rachis, primary and secondary branches and pedicels, are also infected.

Control

Photo by: Claudio Bezerra (BRSMG Curinga lineages that were edited and inoculated with Magnaporthe oryzae to check for resistance to blast)

Currently, damage caused by blast can be reduced by integrating the use of resistant cultivars, crop practices and fungicides to crop management with suitable soil preparation; balanced fertilization, avoiding excessive vegetative plant growth; use of seeds of good phytosanitary and physiological quality; sowing in a minimal period of time and in the opposite direction of the predominant wind direction; incorporation of crop remains; uniform sowing depth; recommended seeding density for the cultivar or production system.

Other important practices include weed control; destruction of volunteer and diseased plants; good soil leveling; maintaining the appropriate irrigation water level during the plant cycle; adequate dimensioning of irrigation and drainage systems; change of choice of cultivars every 3 or 4 years; sowing at the beginning of the rainy season; use of fungicides applied in seed treatment and spraying of aerial plant parts. 

Protection against panicle blast is performed preventively through spraying with systemic fungicides: one application should take place at the end of the rubberization period and the other one when there is up to 5% panicle emission.

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