In the segment 'A Look Around Innovation,' we introduce research and development (R&D) sites that have led to social implementation. In the 16th installment of this series, we introduce the efforts of Professor Tamao Saito of the Faculty of Science and Technology at Sophia University, who is collaborating with a company to develop plant protection materials from nematode repellents produced by "cellular slime molds," which have characteristics of both animals and plants to reduce the pesticide use.
Only one previously published study: Experiments with root-knot nematodes
Despite its location near the JR Chuo Line Yotsuya Station in the center of the capital, Sophia University's Yotsuya Campus is surrounded by lush greenery. Professor Tamao Saito of the Faculty of Science and Technology, Sophia University, assumed her position in 2009 and initiated research into the relationship between cellular slime molds and nematodes the following year.
"Cellular slime molds" are soil microorganisms found near the surface of the earth. They are unicellular amoebae, but are a rare in that they use a survival strategy of aggregating and behaving as mushroom-like multicellular organisms when bacteria are not available as prey. Saito learned about cellular slime molds when she was a university student and has continued to research them because she is intrigued by the possibility that "such research might reveal the process of evolution from unicellular organisms to multicellular organisms."
After completing graduate school, she engaged in the genome analysis of cellular slime molds in the genome sequencing project of model organisms that has its roots in the Human Genome Project. The results showed that 62% of the whole cellular slime mold genome consists of coding regions, which serve as blueprints for proteins, and many of them code "polyketide synthases." Polyketides are secondary metabolites. Primary metabolites, such as proteins and carbohydrates, are essential for the vital activities of organisms. Secondary metabolites serve functions to survive in the natural world and are unique to each organism. Based on their functions, these substances are used as pharmaceutical products and their raw materials. Additionally, some polyketides are used to communicate with other organisms.
"After I wondered about what cellular slime molds are doing with polyketide synthases, I began to think that they could be used to communicate with other microorganisms for survival," Saito said. At the same time, she noticed that nematodes were always present in soils in which cellular slime molds were found. Saito thought that there must be some ecological relationship between the two and began her research. The only previous study was a paper published in 1996 by overseas researchers. The movements of nematodes placed in the center of a petri dish were examined in the presence of the slime mold amoebae or food on one side of the petri dish. The nematodes moved freely in the petri dish when neither slime mold nor food was placed, and the nematodes moved toward the food when only food was placed. However, when only the slime mold amoebae were placed without food, the nematodes moved away from the slime mold amoebae toward the other side.
Based on this paper, Saito conducted the same experiment with the plant-parasitic root-knot nematodes (Meloidogyne spp.) and found that the nematodes moved to avoid the slime mold fruiting bodies. Furthermore, when the cellular slime mold was grown on a piece of filter paper in a petri dish, and the nematodes were placed in the dish after the slime mold and filter paper were removed en bloc, the nematodes still avoided where the slime mold was (Figure 1). These results suggest that some chemical compounds secreted by slime molds also act to repulse nematodes. Saito thought that keeping nematodes away was advantageous for the survival of slime mold, and she had a sudden flash that said this could be used for agricultural purposes.
Figure 1:Relationship between cellular slime molds and nematodes
From a New Technology Presentation Meeting to joint research — Mass production and active ingredient identification the keys
The root-knot nematodes used in the experiment parasitize many crops and form "root-not galls," causing serious crop loss (Figure 2). Generally, pesticides are applied to the soil before cropping to eradicate nematodes, but nematodes deep underground often survive. Since pesticides cannot be used during cropping, nothing can be done if the plants are parasitized by nematodes that survived the pesticide treatment. "Slime mold-derived plant protection materials could be useful to protect crops from nematodes." Based on this idea, Saito filed a patent application in 2014 with the support of the Sophia University Center for Research Promotion and Support.
Figure 2:Damage caused by root-knot nematodes
With the Center's recommendation, Saito attended JST's new technology presentation meeting in 2014. From this, several companies interested in the research contacted her. One such company was Panefri Industrial Company (Nagaokakyo City, Kyoto Prefecture), which developed a nematode repellant made from plant materials. Because of the similarity in research direction and the company's extensive and reliable know-how on nematode control, Saito decided to collaborate with Panefri. "I have been working mainly on basic research until now, so I didn't realistically project that my research would lead to social implementation. I was very happy to have this valuable opportunity."
In the joint research, Saito was responsible for identifying repellent substances, clarifying their mechanisms of action, and developing mass production technology. Panefri was in charge of mass production for commercialization and verification of nematode control effects in actual fields. The joint research was selected for JST's Matching Planner Program in 2015 and the A-STEP Industry-Academia Collaboration Phase (Seeds development type FS) in 2017. At the beginning of the research, she was often perplexed by the difference in scale.
Until then, Saito had handled only 10-20 milliliters of repellents in her laboratory, but the company requested that she provide several dozen times that scale. Having acknowledged that establishing the mass production system and identifying repellant substances were essential for practical application, Saito discussed this matter with Panefri. After completing the Seeds development type FS, Saito returned to the functional verification phase of A-STEP to solve these two issues before the next industry-academia collaboration (full-scale type) stage.
Path to low-cost metabolite production and aiming to improve soil health in actual fields
Establishing the mass production system required a series of trial and error. Tank culture was thought to be unsuitable for cellular slime molds. However, Saito overcame this problem by adjusting various conditions, such as agitation speed and temperature, and succeeded in dramatically increasing the culture scale. All water-soluble slime mold components with repellent activity were analyzed, and approximately 150 compounds, including analogs, were tested for the nematode-repellent effect.
After identifying 14 different compounds with the effects, she tested a mixture of 9 selected compounds for the repellant activity. The test found that the slime mold extract and the mixture showed almost the same activity when the amount of the latter used was about one-hundredth of the former. This result suggests that a production cost lower than the previous cost may be achievable by mixing the identified repellent ingredients.
The next step toward practical application is to verify the effectiveness in actual fields. Unlike laboratory experiments, the verification in actual fields is inevitably susceptible to the effects of other microorganisms and climate conditions and is associated with increased difficulty. Sufficient results have not yet been obtained in the field, but she will continue the verification study in parallel with analysis of the mechanism. "During our research, we have found that this repellent also has a growth-promoting effect on plants. If we can put it to practical use, we believe it will reduce the use of chemical pesticides and improve soil health," Saito told us about her future prospects (Figure 3).
Figure 3:Changes expected from the use of nematode repellent materials in agriculture
The Ministry of Agriculture, Forestry, and Fisheries formulated the "MIDORI Strategy for Sustainable Food Systems" in 2022, aiming to improve the productivity and sustainability of the food, agriculture, forestry, and fisheries industries through innovation. The Strategy aims to reduce the use of chemical pesticides by 50% by 2050. This nematode-repellent material, based on research and development on the properties of slime molds by Saito and Panefri, must be one of the effective tools to achieve this goal without burdening producers.
(Article: Yuko Sakurai, Photography: Hideki Ishihara)