Background

Rice is one of the staple foods of 1.4 billion people in our country. Rice production accounts for nearly 40% of our food production. It can be said that rice has provided a livelihood for millions upon millions of families in our country. In the thousands of years of spring planting and autumn harvesting, the Chinese nation has formed a profound and long-standing rice culture, which has spread to all parts of the world and influenced people.

Figure 1. The rice

China is the country with the earliest history of rice cultivation in the world. Modern archaeological studies show that the ancestors in the southeast coastal areas of China began to cultivate rice for survival and breeding during the Neolithic period 12,000 to 7,000 years ago. Along with the migration of population, the rice-based culture blended with the dryland culture in the northern region of the Yellow River basin, and together gave birth to the splendid Chinese civilization.

Figure 2. Ancient Picture: The Picture of Ploughing and Weaving

Since ancient times, many Chinese characters, words and idioms are related to rice. The art of painting, calligraphy and poetry related to rice has enriched and spread people's spiritual world and enriched Chinese culture. What's more, rice can be made into a variety of rice food with rich taste to feed a huge population. The long planting culture and rich food culture such as glutinous rice flour, glutinous rice cake, glutinous rice balls and rice wine show that Chinese people have a deep and inseparable connection with rice.

"The Silk Road is for the aristocracy, the rice road is for the masses."

-- The Rice Road

Overview

Severity of Rice Sheath Blight

With over three billion people across the globe eating rice every day as their stable food, rice is critical to global food security, thus, ensuring sustainable rice production is a key contribution to the global goal of ending hunger. However, the production of rice is affected by the global climate as well as pests and pathogens, which contain Rice Sheath Blight(ShB), a severe rice damage disease caused by Rhizoctonia solani AG1-IA(R.solani). The pathogen causes rice yield damage to 45% and causes damage to rice grain filling and yield in rice. Otherwise, R.solani has a wide range of host crops such as potato, wheat and soybean, which are both crucial food crops and economic crops all around the world.

In China, ShB was first reported in 1934, and it has become the second most important disease in rice at present, causing a yield loss of 10~30% every year, even up to 50% in the rice-growing region of Yangtze river valley and South China in epidemic years.The annual disease area in China is about 15~20 million `hm^2`(2019). Figure 3 shows the host crops of R.solani and its global distribution.

Figure 3. The global distribution of R.solani.

The Defects of Existing Methods

The pathogenic fungus of ShB, R.solani, is a soil-borne fungus that exists as a sclerotia when the environment is not conducive to growth, such as in soil. The sclerotia is a solid dormant mass of tangled mycelium and is considered to be the main source of infection for blight. The compact structure of the sclerotia and the black pigment in the cell wall protect the sclerotia from external influences such as UV light.

When planting rice, the process of irrigating the field after planting rice seedlings will cause the sclerotia present in the soil to float on the water surface, and the floating sclerotia will flow through the aquatic interface and infect other rice plants. Meanwhile, R.solani prefers high temperature and high humidity, 28°C-32°C is its optimum growth temperature. The high temperature and high humidity environment of planting rice is conducive to the germination of sclerotia, and the growing mycelium will spread laterally to infect other rice plants.

Figure 4. Schematic diagram of R.solani infection process in rice.

Rice sheath blight can occur throughout the entire reproductive cycle of rice, and is generally most severe from the beginning of the tillering stage to around the time of tasseling. It mainly affects leaf sheaths and leaves, but in severe cases it can also affect stalks and spikes. Therefore, it is important to control the spread of the sclerotia and R.solani. Early detection in the field is essential to prevent the spread of sclerotia at an early stage. Traditional methods rely on visual recognition by experienced farmers, but by this time the sclerotia has already spread throughout the field and disease spots on leaves are present, meaning that the rice is already infected.

Figure 5. (a)The dark brown particles are sclerotia of R.solani. (b)This picture shows rice leaves infected by R.solani.

With the advance of technology, spectral detection methods such as using hyperspectral images have been tested for detection disease spots. However, this requires personnel and it is large and expensive.

In addition, the existing means of control of R.solani and rice sheath blight are also facing difficulties. The traditional physical salvage of the sclerotia, which requires a large area to be salvaged each season and taken out of the field for deep burial, is time-consuming and labour-intensive. The chemical pesticides commonly used by farmers also face problems of drug resistance, and this was echoed in our interviews with farmers in our human practices.

At the same time, due to the lack of resistant germplasms in rice, progress in breeding for ShB-resistant varieties is slow. Additionally, it is also difficult to screen for resistant species of other crops, such as soybean, a crop widely grown in North America and of high economic value.

The developing resistance to existing chemical pesticides and the complexity and difficulty of screening for resistant genes due to a wide range of hosts have led to the continued worldwide prevalence of plant diseases caused by R.solani.

Table 1. Host range of R.solani.

Policy Support

Biopesticide

In August 2021, China announced the "14 · Five" National Agricultural Green Development Plan. The article in Chapter 4, Section 2 proposed the implementation of green prevention and control, in key areas of horticultural crops, integrated promotion of biological control, physical control and other green prevention and control technologies. The policy wants to promote the concentration of production factors toward bio-derived pesticides, and promote the transformation and upgrading of the pesticide industry and its sustainable and healthy development.

At the same time, in order to encourage enterprises' technological innovation and research and development, China has issued a series of preferential tax policies. All qualified leading enterprises of bio-derived pesticides can enjoy preferential tax policies according to regulations. Under the premise of ensuring safety and effectiveness, the Ministry of Agriculture and Rural Affairs has reduced the test content and shortened the test period according to the characteristics of different types of biogenic pesticides, and encouraged enterprises to develop and register high-efficiency and low-risk biogenic pesticides.

At present, global organic agriculture is developing rapidly, residents' awareness of environmental protection is increasing, and policies vigorously support the development of biopesticide industry. According to IHS Markit, the biopesticide industry was valued at about $5 billion in 2020 and will continue to grow rapidly. The global biopesticide industry market size will grow at a CAGR of approximately 10% during 2020-2025 to reach an estimated size of over US $8 Billion by 2025. Based on this growth rate, the global biopesticide industry market size is expected to reach approximately US $8.8 billion in 2026.

Figure 6. Forecast of global biopesticide industry market prospect from 2021-2026 (unit: 100 million dollars).

Precision Agriculture/Digital Agriculture

Current FAO projections indicate that the global population could increase by 2.3 billion people from today’s levels, reaching 9.8 billion by 2050. As population growth and environmental pressures become issues facing the global food system, digital agriculture and precision agriculture have been proposed as a means to address these issues. They can help "optimize crop growth and yields" (World Bank, 2017, p. 108),"optimize the utilization of natural resources" (FAO, 2018a, p. 25).

In 2022, China issued the "Implementation Plan for the In-depth Promotion of agricultural digital Construction in the 14th Five-Year Plan" to accelerate the construction and application of big data in agriculture and rural areas, focus on the construction of the national agricultural and rural big data platform, establish and improve the agricultural and rural data resources system, and build the national agricultural and rural big data "one map". Among the 14 key initiatives, Article 6 refers to the digitalization of capacity monitoring and Article 8 refers to the digitalization of productive services.

RiceAide

As mentioned above, there is almost no effective scientific means to prevent and control rice sheath blight, which causes serious losses to paddy fields and troubles the vast number of people who live on rice. This year, we were inspired by the trend of digital agriculture and the development of new control methods in biology academia. As students in a rice-growing country, we felt it is our responsibility to improve this situation, make breakthroughs and combine various technologies to treat rice sheath blight. Therefore, we created "RiceAide", a comprehensive control project for rice sheath blight based on the "prevention-detection-treatment" mode. Here comes the introduction of these three steps with their own features.

1.Prevention

"Aquatic interface! Let's block the transmission route of Rhizoctonia solani!"
"Just create 'Trichoderma's acest carrier ever(TACE)' to keep our engineered fungi floating!"

Since rice sheath blight is so common and potentially devastating, farmers need a right-hand assistant to prevent rice sheath blight in daily planting. We brought in one of our main characters, Trichoderma soldiers in a "TACE" boat, to the aquatic interface of rice crops, which is the infection site of R.solani.

Considering cost and sustainability, we apply a mixture of engineered Trichoderma (T.atroviride) and common T.atroviride on a large scale in the field. During this process, engineered T.atroviride will express the hydrophobic protein Epl 1, with the aid of our "TACE" hydrophobic material, will accumulate at the aquatic interface of rice to prevent R.solani from infesting the rice. T.atroviride will also overexpress Prb 1, a type of serine protease playing great importance in mycoparasitism through hydrolyzing the cell wall of R.solani. Lastly, engineered T.atroviride will express Snakin 1, a potato-derived cysteine-rich antimicrobial peptide inhibiting the growth of a variety of plant pathogens, especially R.solani. By these steps, most R.solani can be inhibited and killed.

For safety, we designed a condition-triggered suicide switch, which prevents engineered T.atroviride from escaping and bringing negative effects to the field ecological environment.

2.Detection

"E-nose and LAMP-LFD detection system could help farmers find the pathogen, Rhizoctonia solani, which hides in rice fields and is hard to observe."
"SmartFarm APP can reassure farmers about the growing conditions of rice and provide detection feedback."

We are determined to overcome the lack of routine and early detection methods as well as make contributions to digital agriculture. In addition to preventive measures, we established a daily detection method, the field "lighthouse" - E-nose. And an accurate detection method, the "guard" - LAMP-LFD device.

In terms of detection, we first designed an electronic hardware called ENose. It can collect data such as temperature, humidity, light intensity, wind speed and gas concentration of the surrounding environment through the built-in sensor, and transmit them to the detection algorithm running on our server for data processing and analysis, and finally send the analysis results to users through the APP. When ENose detects an anomaly in the data, it indicates that crops in the rice field may be infected with a disease, and the system will automatically alert users.

After E-nose detected field abnormalities, we used a compact LAMP-LFD detection device, which is portable, fast and sensitive. The LAMP reaction targets at the ITS sequence of R.solani and achieve accurate detection. In addition, the LAMP-LFD detection system can be useful for farmers and scientists when rice sheath blight is high in high temperature and high humidity climate, or after spraying our Trichoderma TACE sustained release tablets and RNAi in the field.

3.Treatment

"RNAi small molecules are easy to degrade, let's use nanomaterials to keep them stable and easy for R.solani to absorb."
"shRNA will be more stable! Let's use E.coli to produce them!"

When E-nose and LAMP-LFD detect rice sheath blight, our "paratrooper" RNAi molecules will land in the appropriate area and kill R. solani. Their "parachutes" are nanomaterials, which makes them less vulnerable to degradation.

RNAi technology, with high specificity and biosafety, is our treatment method to inhibit Rhizoctonia solani. After fermentation by E.coli, we can obtain a large number of shRNA molecules under the cleavage of R-body, then bind them with nanomaterials, and finally obtain our RNAi products. When rice sheath blight is detected, RNAi products produced by us will be sprayed on the affected farmland in a large area by UAV, so as to have a therapeutic effect on rice sheath blight in this area.

Figure 7.RNA biopharmaceutical production process.

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