A method for improving yield and quality of rice by combined application in stages

Abstract

This invention discloses a staged combined application method for improving rice yield and quality. The method includes: during the basal fertilizer application stage, applying slow-release fertilizer, *Trichoderma harzianum* solid inoculant, and superphosphate into the paddy soil. The total nitrogen application rate for the entire rice growth period is 10 kg / mu (pure nitrogen), and all nitrogen is applied in a single application during the basal fertilizer stage using slow-release fertilizer; spraying a compound solution at 30 L / mu during the jointing and booting stages, with the *Trichoderma harzianum* solid inoculant and potassium humate concentrations at 5 g / L and 0.5 g / L respectively, and a salicylic acid concentration of 150 ppm; and supplementing with 2 kg / mu of potassium chloride fertilizer 7–10 days before heading. This invention can increase the number of effective panicles and yield in rice, and promote tillering, dry matter, and carbohydrate accumulation. The combined application of slow-release fertilizer and *Trichoderma harzianum* solid inoculant is beneficial for increasing the head rice rate and reducing chalkiness.

Description

Technical Field

[0001] This invention belongs to the field of rice cultivation and agricultural fertilizer application technology, specifically relating to a method for staged combined application to increase rice yield and improve rice quality. Background Technology

[0002] Rice yield and quality are influenced by a variety of factors, including varietal characteristics, soil nutrient supply, fertilization system, and growth period regulation. Current rice cultivation often employs a multi-stage fertilization approach, combining basal fertilizer, tillering fertilizer, and panicle fertilizer. Multiple applications of fast-acting nitrogen fertilizer increase the number of field operations and labor input. Meanwhile, nitrogen supply in paddy fields is closely related to water management and soil conditions, and there is still room for improvement in the stability of nutrient supply.

[0003] Slow-release fertilizers can release nutrients over a longer period, which helps reduce the frequency of nitrogen fertilizer application and maintain nutrient supply throughout the rice's growth cycle. (Trichoderma harzianum) Trichoderma harzianum Rifai is a commonly used beneficial microorganism in agricultural production. Its inoculants can be used for rhizosphere soil regulation and plant growth promotion. Potassium humate and salicylic acid can be used as exogenous regulatory substances for foliar spraying, participating in the regulation of crop growth, stress resistance, and grain filling processes.

[0004] In existing technologies, slow-release fertilizers, Trichoderma harzianum solid inoculants, potassium humate, and salicylic acid are mostly applied individually or in combination. There is still a lack of a complete set of application procedures that connect the slow-release nutrient supply and rhizosphere microbial regulation at the basal fertilizer stage, the foliar spraying of ternary compound solutions at the jointing and booting stages, and potassium supplementation before heading, all tailored to different growth stages of rice.

[0005] Especially under the condition that nitrogen fertilizers are no longer applied during the rice growth period, how to balance the early rice population building, mid-to-late panicle and grain formation and grain carbohydrate accumulation, so as to steadily increase rice yield and take into account the rice processing quality and appearance quality, still requires further research. Summary of the Invention

[0006] The purpose of this invention is to provide a method for staged combined application of fertilizers to improve rice yield and quality, in order to solve the problems of independent soil basal application and mid-to-late stage foliar regulation in the existing rice fertilization process, and insufficient connection between regulation at different growth stages.

[0007] This invention combines slow-release fertilizer, Trichoderma harzianum solid inoculant, and superphosphate into the paddy soil during the basal fertilizer application stage, ensuring that all nitrogen application during the entire rice growth period is supplied through slow-release fertilizer at the basal fertilizer stage. A compound solution containing Trichoderma harzianum solid inoculant, potassium humate, and salicylic acid is sprayed at the jointing and booting stages, respectively. Simultaneously, potassium chloride fertilizer is applied 7–10 days before heading, forming a phased combined application method that integrates rhizosphere and foliar application, and connects early-stage population building with mid-to-late-stage panicle and grain formation regulation.

[0008] To achieve the above objectives, the present invention adopts the following technical solution: This invention provides a method for staged combined application of pesticides to increase rice yield and improve rice quality, comprising the following steps: (1) Base fertilizer application: During the base fertilizer application stage of rice, slow-release fertilizer, Trichoderma harzianum solid inoculant and superphosphate are applied to the paddy soil; the application rate of the slow-release fertilizer is 40 kg / mu, the nitrogen content of the slow-release fertilizer is 25% by mass, the nitrogen application rate of rice during the whole growth period is 10 kg / mu based on pure nitrogen, all the nitrogen is applied at once through the slow-release fertilizer during the base fertilizer application stage, and no additional nitrogen fertilizer is applied during the rice growth period; the application rate of the Trichoderma harzianum solid inoculant is 1 kg / mu, and the application rate of the superphosphate is 32.5 kg / mu; (2) Foliar spraying: The compound solution was sprayed on the leaves and stems of rice at the jointing stage and the booting stage, respectively. It was sprayed once at each of the two growth stages, and the amount sprayed per mu was 30 L each time. The mass concentrations of Trichoderma harzianum solid inoculant and potassium humate in the compound solution were 5 g / L and 0.5 g / L, respectively, and the concentration of salicylic acid was 150 ppm. (3) Potassium supplementation stage: 7-10 days before rice heading, apply potassium chloride fertilizer evenly to the paddy field at a rate of 2 kg / mu.

[0009] Furthermore, the labeled contents of P2O5 and K2O in the slow-release fertilizer are 12% and 15%, respectively.

[0010] Furthermore, the effective phosphorus content of the superphosphate is 16% (based on P2O5).

[0011] Furthermore, the effective potassium content of the potassium chloride fertilizer is 60% (calculated as K2O).

[0012] Furthermore, the Trichoderma harzianum solid inoculant is a powdered solid inoculant with an effective viable count of not less than 1.0 × 10⁻⁶ cells / year. 9 CFU / g.

[0013] Furthermore, the same Trichoderma harzianum solid inoculant was used for both basal fertilizer application and foliar spraying.

[0014] Furthermore, the rice is conventional indica rice or conventional japonica rice.

[0015] In this invention, the labeled nutrient content of slow-release fertilizer is calculated as N-P2O5-K2O; the available phosphorus content of superphosphate is calculated as P2O5; and the available potassium content of potassium chloride fertilizer is calculated as K2O. Unless otherwise stated, the relevant nutrient content in the fertilizer is all by mass percentage; the concentrations of Trichoderma harzianum solid inoculant and potassium humate in the compound solution are all by mass concentrations, and the concentration of salicylic acid is expressed in ppm.

[0016] In this invention, *Trichoderma harzianum* solid inoculum refers to a commercially available powdered *Trichoderma harzianum* solid inoculum with an effective viable count of not less than 1.0 × 10⁻⁶ cells / year. 9 CFU / g. The same Trichoderma harzianum solid inoculant is used for both basal fertilizer application and foliar spraying, only the application method, application rate or concentration is different: the application rate is 1 kg / mu during the basal fertilizer application stage, and the mass concentration in the compound solution for foliar spraying is 5 g / L.

[0017] Compared with the prior art, the present invention has at least the following beneficial effects.

[0018] (1) This invention combines slow-release fertilizer, Trichoderma harzianum solid inoculant and superphosphate as basal application with spraying Trichoderma harzianum solid inoculant, potassium humate and salicylic acid compound solution during the jointing and booting stages, and supplements potassium chloride fertilizer before heading, forming a staged combined application method adapted to different growth stages of rice.

[0019] (2) Under the condition that the nitrogen application rate of rice during the whole growth period is 10 kg / mu of pure nitrogen, the present invention enables all nitrogen to be applied through slow-release fertilizer at the basal fertilizer stage, and no nitrogen fertilizer is applied during the rice growth period, which can reduce the number of nitrogen fertilizer applications and simplify field management.

[0020] (3) The application of slow-release fertilizer in combination with Trichoderma harzianum solid inoculant can increase rice yield and improve rice quality. The experimental results showed that this treatment increased the yield of Huanghuazhan and Jia58 by 7.3% and 9.9% respectively compared with conventional fertilization, and Huanghuazhan rice showed a higher head rice rate and lower chalkiness.

[0021] (4) The compound solution used in this invention is composed of Trichoderma harzianum solid inoculant, potassium humate and salicylic acid. After spraying during the jointing and booting stages, the average yields of Jiayou No. 5 and Huazheyou 210 increased by 8.2% and 9.7% respectively compared with the control in clear water, indicating that the compound solution has a good yield-increasing effect.

[0022] (5) After adopting the phased combined application method of the present invention, the yield of Huanghuazhan and Longjing 31 increased by 16.5% and 19.9% ​​respectively compared with conventional fertilization, and the number of effective panicles increased significantly. The number of grains per panicle, the seed setting rate and the weight of a thousand grains showed different degrees of increase.

[0023] (6) The present invention can promote the occurrence of rice tillers, the retention of stem tillers, the accumulation of dry matter and carbohydrates; among them, the activity of ADPG pyrophosphorylase in Huanghuazhan grains is significantly increased, and the activities of soluble starch synthase and starch branching enzyme show an increasing trend, which is beneficial to the formation of rice yield. Attached Figure Description

[0024] Figure 1 The effects of different basal application treatments on rice appearance, head rice yield, and chalkiness are shown, where a represents the appearance of polished rice, b represents the head rice yield, and c represents the chalkiness. Different lowercase letters indicate the differences between treatments. P The difference is significant at the level of <0.05. The same lowercase letter indicates that the difference between treatments is not significant.

[0025] Figure 2 The effects of Trichoderma harzianum application on rice plant morphology and panicle morphology were investigated, where a represents rice plant morphology and b represents panicle morphology.

[0026] Figure 3 The effects of Trichoderma harzianum application on rice plant height and tiller dynamics were investigated, where a represents the change in rice plant height and b represents the change in the number of rice tillers.

[0027] Figure 4 The effects of Trichoderma harzianum application treatments on the activities of rice grain-filling related enzymes were investigated. In this study, a represents ADPG pyrophosphorylase activity, b represents soluble starch synthase activity, and c represents starch branching enzyme activity. Different lowercase letters indicate differences in treatment activity. P A difference of <0.05 is considered significant. The same lowercase letter or the presence of the same lowercase letter indicates that the difference between treatments did not reach a significant level. Detailed Implementation

[0028] The present invention will be further described below with reference to specific embodiments. It should be understood that the following embodiments are for illustrative purposes only and are not intended to limit the scope of protection of the present invention.

[0029] Unless otherwise stated, the slow-release fertilizer, superphosphate, potassium chloride fertilizer, Trichoderma harzianum solid inoculant, potassium humate and salicylic acid used in the following examples are all conventional commercially available products.

[0030] The labeled nutrient content of slow-release fertilizer is 25%-12%-15% (N-P2O5-K2O); the available phosphorus content of superphosphate is 16% (P2O5); and the available potassium content of potassium chloride fertilizer is 60% (K2O).

[0031] Trichoderma harzianum solid inoculant is a commercially available powdered solid inoculant, with a product specification of 1 kg / bag and an effective viable count of not less than 1.0 × 10⁹ CFU / g. The same Trichoderma harzianum solid inoculant is used for both basal fertilizer application and foliar spraying; the application rate during basal fertilizer application is 1 kg / mu, and the mass concentration in the compound solution for foliar spraying is 5 g / L.

[0032] The compound solution is prepared by adding water to Trichoderma harzianum solid inoculum, potassium humate, and salicylic acid. The Trichoderma harzianum solid inoculum, potassium humate, and salicylic acid are added to water and mixed thoroughly to obtain the compound solution for foliar spraying. The mass concentrations of Trichoderma harzianum solid inoculum and potassium humate in the compound solution are 5 g / L and 0.5 g / L, respectively, and the concentration of salicylic acid is 150 ppm. Based on 30 L of the compound solution per acre per spray, each spray contains 150 g of Trichoderma harzianum solid inoculum and 15 g of potassium humate; it is sprayed once each at the rice jointing stage and the booting stage.

[0033] The nutrient input of each fertilizer in this invention is calculated based on N, P2O5, and K2O, as follows: Slow-release fertilizer 40 kg / mu, of which the nitrogen content is 25% by mass, providing 10 kg / mu of N; the slow-release fertilizer has a labeled P2O5 content of 12%, providing 4.8 kg / mu of P2O5; superphosphate 32.5 kg / mu, with an available phosphorus content of 16% (based on P2O5), providing 5.2 kg / mu of P2O5, for a total of 10 kg / mu of P2O5; the slow-release fertilizer has a labeled K2O content of 15%, providing 6 kg / mu of K2O; potassium chloride fertilizer 2 kg / mu, with an available potassium content of 60% (based on K2O), providing 1.2 kg / mu of K2O, for a total of 7.2 kg / mu of K2O.

[0034] Rice yield was calculated in kg / mu (unit of land area) after harvest from each plot. After three months of storage, the head rice rate and chalkiness were determined according to GB / T 17891-2017 "High-Quality Rice" and its referenced methods. Plant height and tiller number were measured by selecting 10 hills per plot and recording these measurements every 7 days starting one week after transplanting. Dry matter was measured by randomly selecting 10 plant samples from each plot one day before harvest at the jointing, booting, grain-filling, and maturity stages. The samples were separated into leaves, stem sheaths, and panicles. Each sample was placed in an oven, blanched at 105℃ for 30 min, and then dried at 75℃ to constant weight before weighing. Soluble total sugar and starch were determined by the anthrone colorimetric method; the content of non-structural carbohydrates was calculated as the sum of soluble total sugar content and starch content; the activities of ADPG pyrophosphorylase, soluble starch synthase and starch branching enzyme were detected by an ELISA kit (Shanghai Enzyme-Linked Biotechnology Co., Ltd., China).

[0035] Data from each treatment were statistically analyzed using analysis of variance. For tables and bar charts labeled with letters, different lowercase letters indicate differences between treatments. P The difference is significant at the level of <0.05. The same letter indicates that the difference between treatments is not significant.

[0036] Example 1: Effects of combined application of slow-release fertilizer and Trichoderma harzianum solid inoculant on rice yield and rice quality. The experiment was conducted from May to October 2024 in Yangqingmiao Village, Fuyang District, Hangzhou City, Zhejiang Province. The rice varieties tested were Huanghuazhan, a conventional indica rice variety, and Jia 58, a conventional japonica rice variety.

[0037] The experiment employed a randomized block design, with three replicates for each treatment. Rice was sown around May 20th and transplanted from June 11th to 12th, using double-stem planting with a plant spacing of 20 cm × 25 cm. Each plot had an area of ​​20 m². 2 .

[0038] Configure the following process: NF: No fertilizer application treatment; CF: Conventional fertilization treatment; SRF: Slow-release fertilizer treatment; SRFT: Treatment involving the combined application of slow-release fertilizer and Trichoderma harzianum solid inoculant; SRFT-N10%: Treatment involving the application of slow-release fertilizer (10% nitrogen reduction) and Trichoderma harzianum solid inoculant; SRFT-N20%: Treatment involving the application of slow-release fertilizer (nitrogen reduction of 20%) and Trichoderma harzianum solid inoculant.

[0039] Except for the NF treatment, the nutrient application rates for rice throughout its entire growth period were determined according to the experimental design. The total nitrogen application rate for the conventional fertilization treatment and the equal nitrogen slow-release fertilizer treatment was equivalent to 10 kg / mu of pure nitrogen, the phosphorus application rate was equivalent to 10 kg / mu of phosphorus pentoxide, and the potassium application rate was equivalent to 7.2 kg / mu of potassium oxide.

[0040] The slow-release fertilizer used in this embodiment has a labeled nutrient content of 25%-12%-15% (N-P2O5-K2O); the effective phosphorus content of the superphosphate fertilizer is 16% (P2O5); and the effective potassium content of the potassium chloride fertilizer is 60% (K2O). The specific fertilization methods and application rates for each treatment are shown in Table 1.

[0041] Table 1. Fertilizer application rates at different fertilization stages for different treatments

[0042] Note: NF, no fertilization; CF, conventional fertilization; SRF, slow-release fertilizer; SRFT, slow-release fertilizer combined with Trichoderma harzianum solid inoculant; SRFT-N10%, slow-release fertilizer with 10% nitrogen reduction and combined with Trichoderma harzianum solid inoculant; SRFT-N20%, slow-release fertilizer with 20% nitrogen reduction and combined with Trichoderma harzianum solid inoculant. Potassium chloride in the table refers to potassium chloride fertilizer, with an effective potassium content of 60% (K2O).

[0043] The actual yield of each treatment was measured after the rice matured, and the results are shown in Table 2.

[0044] Table 2 Actual yield of rice under different fertilization treatments

[0045] Note: Yield increase 1 represents the yield change of each treatment compared to the NF (no fertilization) treatment, and yield increase 2 represents the yield change of each treatment compared to the CF (conventional fertilization) treatment. For the same variety in the same column, different labels without identical lowercase letters indicate significant differences between treatments at the P < 0.05 level; labels with identical lowercase letters or containing identical lowercase letters indicate that the differences between treatments did not reach a significant level.

[0046] As shown in Table 2, different fertilization treatments had different effects on the yield of the two rice varieties.

[0047] For Huang Huazhan, the average yields of the NF, CF, SRF, SRFT, SRFT-N10%, and SRFT-N20% treatments were 395.9 kg / mu, 513.1 kg / mu, 519.5 kg / mu, 550.8 kg / mu, 522.5 kg / mu, and 507.3 kg / mu, respectively. The SRFT treatment had the highest average yield, increasing by 7.3% compared to the CF treatment and by 39.1% compared to the NF treatment.

[0048] For Jia 58, the average yields of the NF, CF, SRF, SRFT, SRFT-N10%, and SRFT-N20% treatments were 400.5 kg / mu, 501.7 kg / mu, 507.3 kg / mu, 551.2 kg / mu, 510.1 kg / mu, and 468.7 kg / mu, respectively. The SRFT treatment had the highest average yield, increasing by 9.9% compared to the CF treatment and by 37.6% compared to the NF treatment.

[0049] This indicates that, under the same nitrogen application conditions, the combination of slow-release fertilizer and Trichoderma harzianum solid inoculant can increase the yield of Huanghuazhan and Jia 58 rice varieties.

[0050] Under the SRFT-N10% treatment, the yields of Huanghuazhan and Jia58 rice varieties increased by 1.8% and 1.7% respectively compared to the CF treatment, indicating that under the conditions of this experiment, when the nitrogen content of slow-release fertilizer was reduced by 10% and combined with the application of Trichoderma harzianum solid inoculant, rice yield could be maintained at a level comparable to or slightly higher than that of conventional fertilization. The yield of the SRFT-N20% treatment was lower than that of the CF treatment, indicating that further reduction in nitrogen fertilizer application was not conducive to maintaining rice yield.

[0051] Further measurements were performed on the appearance, head rice rate, and chalkiness of rice harvested from Huang Huazhan's various treatments. The results are shown in the table below. Figure 1 .

[0052] like Figure 1 As shown in a, different fertilization treatments have a certain impact on the appearance of polished rice.

[0053] like Figure 1 As shown in b, the average head rice percentage of each treatment, from highest to lowest, is SRFT, SRF, SRFT-N10%, CF, SRFT-N20%, and NF. The head rice percentage of each fertilization treatment is higher than that of the NF treatment, with the SRFT treatment having the highest head rice percentage. This indicates that the application of slow-release fertilizer combined with Trichoderma harzianum solid inoculant is beneficial for improving the quality of rice processing.

[0054] like Figure 1 As shown in Figure c, the trend of chalkiness in each treatment is opposite to that of the overall head rice yield. The NF treatment has the highest chalkiness, the SRFT treatment has the lowest chalkiness, and the chalkiness of the other fertilization treatments falls between the two. This indicates that slow-release fertilizer combined with Trichoderma harzianum solid inoculant can reduce rice chalkiness and improve the appearance quality of rice.

[0055] In summary, the yields of both tested rice varieties reached their highest levels under the SRFT treatment; meanwhile, Huanghuazhan rice exhibited a higher head rice rate and lower chalkiness. Therefore, the combined application of slow-release fertilizer and Trichoderma harzianum solid inoculant can increase rice yield and improve rice processing quality and appearance.

[0056] Example 2: Effects of different foliar spray components, concentrations, and their combinations on rice yield This embodiment was conducted in Yangqingmiao Village, Fuyang District, Hangzhou City, Zhejiang Province from May to October 2024.

[0057] The rice varieties tested were Zhongzheyou 8, Huazheyou 210, and Jiayou 5, which are suitable for planting in the middle and lower reaches of the Yangtze River. Among them, Zhongzheyou 8 and Huazheyou 210 are indica three-line hybrid rice, while Jiayou 5 is japonica three-line hybrid rice.

[0058] The experiment adopted a randomized block design in the field. Foliar spraying was carried out at the jointing stage and the booting stage of rice, and sprayed once at each stage. Each time, 30 L of spray solution was applied per acre. The spray solution was evenly sprayed on the rice leaves and stems.

[0059] 2.1 Screening of single-component spray concentration The following spraying treatments are set up respectively: (1) Trichoderma harzianum solid inoculum, with mass concentrations of 0.5 g / L, 1 g / L, 5 g / L and 10 g / L respectively; (2) Potassium humate, with mass concentrations of 0.2 g / L, 0.5 g / L and 1 g / L, respectively; (3) Salicylic acid, concentration 150 ppm; (4) Water control.

[0060] Trichoderma harzianum solid inoculant is a powdered solid inoculant with an effective viable count of not less than 1.0 × 10⁻⁶ cells. 9 CFU / g.

[0061] Using Zhongzheyou 8 as the test variety, the yield of rice under different single-component and different concentration spraying treatments was determined. The results are shown in Table 3.

[0062] Table 3. Effects of different foliar spraying components and concentrations on rice yield.

[0063] The results showed that after spraying with Trichoderma harzianum solid inoculant, potassium humate, or salicylic acid, the average yield of each treatment increased to varying degrees compared with the water control, with the effect varying depending on the type of component and the spraying concentration. Specifically, when the Trichoderma harzianum solid inoculant was sprayed at concentrations of 1 g / L and 5 g / L, the average rice yield increased by 3.91% and 4.58% compared with the water control, respectively; when the potassium humate was sprayed at a concentration of 0.5 g / L, the average rice yield increased by 4.34% compared with the water control; and when the salicylic acid was sprayed at 150 ppm, the average rice yield increased by approximately 3.97% compared with the water control. In addition, under low concentrations of Trichoderma harzianum solid inoculant (0.5 g / L and 10 g / L), and low concentrations of potassium humate (0.2 g / L and 1 g / L), the average rice yield increased by 1.67%, 0.90%, 2.06%, and 1.30% compared with the water control, respectively. Since the treatments in Table 3 are labeled with the same letter, it indicates that the differences between the single-component treatments and the water control were not statistically significant.

[0064] 2.2 Comparison of different compound treatments Based on the single-component concentration screening results above, a compound experiment was conducted using 5 g / L Trichoderma harzianum solid inoculum, 0.5 g / L potassium humate, and 150 ppm salicylic acid, with the following treatments: (1) Water control; (2) Trichoderma harzianum solid inoculum 5 g / L and salicylic acid 150 ppm; (3) Potassium humate 0.5 g / L and salicylic acid 150 ppm; (4) Trichoderma harzianum solid inoculum 5 g / L, potassium humate 0.5 g / L and salicylic acid 150 ppm.

[0065] The effects of different compound treatments on the yield of Jiayou 5 and Huazheyou 210 are shown in Table 4.

[0066] Table 4. Effects of different foliar compound treatments on rice yield

[0067] Note: The average yield of the same variety under different treatments is indicated by different lowercase letters, which means that the difference between treatments is significant at the P < 0.05 level; the same letter or the presence of the same letter indicates that the difference between treatments is not significant.

[0068] As shown in Table 4, for Jiayou No. 5, the average yields of the water control, the binary compound treatment of Trichoderma harzianum solid inoculant and salicylic acid, the binary compound treatment of potassium humate and salicylic acid, and the ternary compound treatment were 626.9 kg / mu, 658.5 kg / mu, 656.5 kg / mu, and 678.6 kg / mu, respectively.

[0069] The ternary compound treatment had the highest average yield, increasing by 8.2% compared to the water control. The ternary compound treatment was significantly higher than the water control, but the differences between it and the two binary compound treatments were not statistically significant.

[0070] For Huazheyou 210, the average yields of the four treatments were 653.8 kg / mu, 690.6 kg / mu, 687.5 kg / mu and 717.2 kg / mu, respectively.

[0071] The ternary compound treatment had the highest average yield, increasing by 9.7% compared to the water control. The ternary compound treatment was significantly higher than the water control and the binary compound treatment of potassium humate and salicylic acid, but the difference between it and the binary compound treatment of Trichoderma harzianum solid inoculum and salicylic acid was not statistically significant.

[0072] The above results indicate that the ternary compound solution composed of 5 g / L Trichoderma harzianum solid inoculant, 0.5 g / L potassium humate, and 150 ppm salicylic acid achieved the highest average yield in both tested rice varieties, demonstrating a good yield-increasing effect.

[0073] Example 3: Effects of a phased combined application method on rice growth, development, and yield formation. This embodiment was conducted from May to October 2025 in Yangqingmiao Village, Fuyang District, Hangzhou City, Zhejiang Province, and the Northern Rice Research Center of China National Rice Research Institute, Baoqing County, Shuangyashan City, Heilongjiang Province.

[0074] Huanghuazhan (Fuyang, Hangzhou) and Longjing 31 (Baoqing County, Heilongjiang), which are suitable for local planting, were selected as the rice varieties for testing. Huanghuazhan is a conventional indica rice and Longjing 31 is a conventional japonica rice.

[0075] The experiment used a field randomized block design, with the following two treatments: (1) Conventional fertilization treatment; (2) Trichoderma harzianum treatment.

[0076] In this embodiment, the Trichoderma harzianum combined application treatment is the abbreviation of the phased combined application treatment of the present invention in the test results. Specifically, it refers to the treatment of applying slow-release fertilizer, Trichoderma harzianum solid inoculant and superphosphate during the basal fertilizer application stage, spraying a compound solution containing Trichoderma harzianum solid inoculant, potassium humate and salicylic acid during the jointing and booting stages of rice, and applying potassium chloride fertilizer during the panicle fertilizer supplementation stage.

[0077] Each treatment was replicated in triplicate. Rice was sown around May 18 and transplanted from June 10 to 11, using a double-stem planting method with a plant spacing of 20 cm × 25 cm. Each plot was 20 m² in size. 2 .

[0078] The nutrient application rates for the conventional fertilization treatment throughout the entire growth period were equivalent to 10 kg / mu for nitrogen, 10 kg / mu for phosphorus pentoxide, and 7.2 kg / mu for potassium oxide. The specific fertilization method was the same as that for the CF treatment in Example 1.

[0079] Trichoderma harzianum application treatment should be carried out in the following manner: (1) During the basal fertilizer application stage, apply 40 kg / mu of slow-release fertilizer, 1 kg / mu of Trichoderma harzianum solid inoculant, and 32.5 kg / mu of superphosphate to the paddy soil. The labeled nutrient content of the slow-release fertilizer is 25%-12%-15% based on N-P2O5-K2O, and the available phosphorus content of superphosphate is 16% based on P2O5.

[0080] (2) The compound solution was sprayed on the leaves and stems of rice at the jointing stage and the booting stage, respectively, once at each stage, with a spraying amount of 30 L per mu each time. The mass concentrations of Trichoderma harzianum solid inoculant and potassium humate in the compound solution were 5 g / L and 0.5 g / L, respectively, and the concentration of salicylic acid was 150 ppm.

[0081] (3) 7 to 10 days before the rice heads emerge, apply potassium chloride fertilizer evenly to the paddy field at a rate of 2 kg / mu; the effective potassium content of potassium chloride fertilizer is 60% calculated as K2O.

[0082] The total nutrient application rates for the Trichoderma harzianum treatment throughout the rice's growth period were equivalent to 10 kg / mu for N, 10 kg / mu for P2O5, and 7.2 kg / mu for K2O. All nitrogen was applied as slow-release fertilizer during the basal fertilizer application stage, and no additional nitrogen fertilizer was applied during the rice's growth period.

[0083] 3.1 Effects of Trichoderma harzianum combined application on rice yield composition Using conventional fertilization as a control, the effective number of panicles, number of grains per panicle, seed setting rate, thousand-grain weight and yield of rice under the Trichoderma harzianum combined application treatment were measured. The results are shown in Table 5.

[0084] Table 5. Effects of Trichoderma harzianum application on rice yield composition.

[0085] Note: In the same column for the same variety, different labels without identical lowercase letters indicate that the differences between treatments are significant at the P < 0.05 level; labels with identical lowercase letters or containing identical lowercase letters indicate that the differences between treatments are not significant.

[0086] As shown in Table 5, compared with conventional fertilization, the Trichoderma harzianum treatment increased the number of effective panicles and yield of the two tested rice varieties.

[0087] The number of effective panicles in Huanghuazhan increased from 202,000 panicles / mu to 219,000 panicles / mu, the number of grains per panicle increased from 147.7 grains to 156.1 grains, the seed setting rate increased from 82.3% to 82.9%, the thousand-grain weight increased from 21.3 g to 21.5 g, and the yield increased from 523.0 kg / mu to 610.0 kg / mu, an increase of approximately 16.5%.

[0088] The number of effective panicles of Longjing 31 increased from 225,000 panicles / mu to 235,000 panicles / mu, the number of grains per panicle increased from 105.6 grains to 117.4 grains, the seed setting rate increased from 93.2% to 94.6%, the thousand-grain weight increased from 22.8 g to 23.2 g, and the yield increased from 504.9 kg / mu to 605.5 kg / mu, representing an increase of 19.9%.

[0089] The above results indicate that, compared with conventional fertilization, the application of Trichoderma harzianum significantly increased the number of effective panicles and yield of Huanghuazhan and Longjing 31 varieties; the number of grains per panicle, seed setting rate and thousand-grain weight of both varieties showed varying degrees of increase.

[0090] 3.2 Effects of Trichoderma harzianum combined treatment on plant morphology, plant height, and tiller dynamics of Huanghuazhan plants The morphology of mature Huanghuazhan plants and rice panicles is shown in [the original text]. Figure 2 a and b in the example.

[0091] like Figure 2 As shown in a, compared with the conventional fertilization treatment, the upper stems and leaves of *Trichoderma harzianum* under the combined treatment were more lush, and the root system was more robust.

[0092] like Figure 2 As shown in b, Huanghuazhan rice under the combined treatment of Trichoderma harzianum showed better panicle development and fuller grain arrangement.

[0093] Rice plant height was measured at 7, 14, 21, 28, 35, 42, 49, and 56 days after transplanting Huang Huazhan rice. The results are shown below. Figure 3 'a' in 'a'.

[0094] like Figure 3 As shown in Figure a, the height of *Trichoderma harzianum* plants in both treatments showed a continuous increasing trend with the number of days after transplanting. At each measurement time, the plant height of the *Trichoderma harzianum* treatment was generally higher than that of the conventional fertilization treatment. At 7 days after transplanting, the difference in plant height between the two treatments was small; however, as the growth progressed, the height advantage of the *Trichoderma harzianum* treatment gradually increased.

[0095] The number of rice tillers was measured simultaneously at each of the above-mentioned measurement times, and the results are shown below. Figure 3 b in the example. Figure 3As shown in b, the number of tillers in both treatments of *Huanghua* showed a trend of first increasing and then decreasing. From 7 to 35 days after transplanting, the number of tillers gradually increased, reaching a relatively high level around 35 days post-transplanting; from 35 to 56 days post-transplanting, the number of tillers gradually decreased.

[0096] During most of the measurement period, the number of tillers in the Trichoderma harzianum treatment was higher than that in the conventional fertilization treatment, especially during the peak tillering period of 28 and 35 days after transplanting, when the number of tillers in the Trichoderma harzianum treatment was significantly higher; in the later stages of growth, the Trichoderma harzianum treatment still maintained a higher number of tillers.

[0097] The above results indicate that the combined application of Trichoderma harzianum can promote tillering of Huanghuazhan and increase the number of tillers retained in the later stages of Huanghuazhan's growth.

[0098] 3.3 Effects of Trichoderma harzianum combined application treatment on dry matter accumulation in rice The stem dry weight, leaf dry weight, panicle dry weight, total plant dry weight, leaf area index, and specific leaf area were measured at the jointing stage, booting stage, grain filling stage, and maturity stage of Huanghuazhan. The results are shown in Table 6.

[0099] Table 6. Effects of Trichoderma harzianum combined treatment on the dry matter accumulation of Huanghuazhan.

[0100] Note: For the same indicator at the same reproductive period, different lowercase letters indicate that the differences between treatments are significant at the P < 0.05 level, while the same lowercase letter indicates that the differences between treatments are not significant.

[0101] As shown in Table 6, during the jointing, booting, grain-filling and maturity stages, the stem dry weight, leaf dry weight, panicle dry weight, total plant dry weight, leaf area index and specific leaf area of ​​Huang Huazhan under the Trichoderma harzianum combined treatment were generally higher than those of the conventional fertilization treatment during the same period.

[0102] At the jointing stage, the total dry weight of the whole plant treated with Trichoderma harzianum was 33.7 g / hole, which was higher than the 25.2 g / hole of the conventional fertilization treatment; the leaf area index was 5.3, which was higher than the 4.1 of the conventional fertilization treatment.

[0103] During the booting stage, the total dry weight of the whole plant treated with Trichoderma harzianum was 45.6 g / hole, which was higher than the 32.9 g / hole of the conventional fertilization treatment; the leaf area index was 7.1, which was higher than the 6.2 of the conventional fertilization treatment.

[0104] During the grouting stage, the total dry weight of the whole plant treated with Trichoderma harzianum was 63.9 g / hole, which was higher than the 45.8 g / hole of the conventional fertilization treatment.

[0105] At maturity, the total dry weight of the whole plant treated with Trichoderma harzianum was 74.5 g / hole, which was higher than the 57.8 g / hole of the conventional fertilization treatment.

[0106] The above results indicate that the combined application of Trichoderma harzianum treatment is beneficial to increasing the leaf area index of Huanghuazhan and promoting the accumulation of dry matter in various organs of Huanghuazhan.

[0107] 3.4 Effects of Trichoderma harzianum combined application treatment on carbohydrate accumulation in rice The contents of soluble total sugar, starch, and non-structural carbohydrates in the stem sheath, leaves, and panicle were measured at the jointing stage, booting stage, grain filling stage, and maturity stage of Huanghuazhan. The results are shown in Table 7.

[0108] Table 7. Effects of Trichoderma harzianum combined treatment on carbohydrate accumulation in Huanghuazhan plants.

[0109] As shown in Table 7, the total soluble sugar content in the stem and sheath of Huanghuazhan rice generally increased with the progress of rice growth; the total soluble sugar content in the leaves showed a trend of first decreasing, then increasing, and then decreasing again; and the starch and non-structural carbohydrate content in the panicle continued to increase from the booting stage to the maturity stage.

[0110] At each measurement period, the total soluble sugar, starch, and non-structural carbohydrate content of Huanghuazhan stems, sheaths, leaves, and spikes under the Trichoderma harzianum treatment were generally higher than those under the conventional fertilization treatment.

[0111] At maturity, the starch content of the ear of Huanghua treated with Trichoderma harzianum was 251.8 mg / g fresh weight, which was higher than that of the conventional fertilization treatment (208.3 mg / g fresh weight); the non-structural carbohydrate content of the ear was 312.1 mg / g fresh weight, which was higher than that of the conventional fertilization treatment (258.9 mg / g fresh weight).

[0112] The above results indicate that the combined application of Trichoderma harzianum treatment is beneficial in increasing the content of soluble total sugar, starch, and non-structural carbohydrates in the stems, sheaths, leaves, and ears of *Trichoderma harzianum*, and promotes the accumulation of carbon matter in the ears during the maturity period.

[0113] 3.5 Effects of Trichoderma harzianum combined application treatment on the activity of enzymes related to starch synthesis in rice grains The activities of ADPG pyrophosphorylase, soluble starch synthase, and starch branching enzyme in Huanghuazhan seeds under different treatments were determined. The results are shown in [Figure number missing]. Figure 4 .

[0114] like Figure 4 As shown in a, the ADPG pyrophosphorylase activity in Huanghuazhan grains under the Trichoderma harzianum combined treatment was significantly higher than that under the conventional fertilization treatment.

[0115] like Figure 4 As shown in b and c, the activities of soluble starch synthase and starch branching enzyme in Huanghuazhan grains under the Trichoderma harzianum combined application treatment were higher than those under the conventional fertilization treatment, but the differences between treatments did not reach a significant level.

[0116] The above results indicate that the combined treatment with Trichoderma harzianum can significantly increase the activity of ADPG pyrophosphorylase in Huanghuazhan grains, and also increase the activities of soluble starch synthase and starch branching enzyme, which is beneficial to starch synthesis and accumulation during the grain filling process of Huanghuazhan grains.

[0117] In summary, this embodiment demonstrates that the combined application of Trichoderma harzianum can increase the effective panicle number and yield of Huanghuazhan and Longjing 31 rice varieties without the need for additional nitrogen fertilizer during the rice growth period. Furthermore, results from Huanghuazhan rice show that this treatment promotes tillering, dry matter accumulation, and carbohydrate accumulation, significantly increases ADPG pyrophosphorylase activity in the grains, and shows an increasing trend in the activities of soluble starch synthase and starch branching enzyme.

Claims

1. A method for staged combined application of pesticides to increase rice yield and improve rice quality, characterized in that, Includes the following steps: (1) Base fertilizer application: During the base fertilizer application stage of rice, slow-release fertilizer, Trichoderma harzianum solid inoculant and superphosphate are applied to the paddy soil; the application rate of the slow-release fertilizer is 40 kg / mu, the nitrogen content of the slow-release fertilizer is 25% by mass, the nitrogen application rate of rice during the whole growth period is 10 kg / mu based on pure nitrogen, all the nitrogen is applied at once through the slow-release fertilizer during the base fertilizer application stage, and no additional nitrogen fertilizer is applied during the rice growth period; the application rate of the Trichoderma harzianum solid inoculant is 1 kg / mu, and the application rate of the superphosphate is 32.5 kg / mu; (2) Foliar spraying: The compound solution was sprayed on the leaves and stems of rice at the jointing stage and the booting stage, respectively. It was sprayed once at each of the two growth stages, and the amount sprayed per mu was 30 L each time. The mass concentrations of Trichoderma harzianum solid inoculant and potassium humate in the compound solution were 5 g / L and 0.5 g / L, respectively, and the concentration of salicylic acid was 150 ppm. (3) Potassium supplementation during panicle: 7-10 days before rice heading, apply potassium chloride fertilizer evenly to the paddy field at a rate of 2 kg / mu.

2. The method according to claim 1, characterized in that, The labeled contents of P2O5 and K2O in the slow-release fertilizer are 12% and 15%, respectively.

3. The method according to claim 1, characterized in that, The effective phosphorus content of the superphosphate is 16% (based on P2O5).

4. The method according to claim 1, characterized in that, The effective potassium content of the potassium chloride fertilizer, calculated as K2O, is 60%.

5. The method according to claim 1, characterized in that, The Trichoderma harzianum solid inoculant is a powdered solid inoculant with an effective viable count of not less than 1.0 × 10⁻⁶ cells / year. 9 CFU / g.

6. The method according to claim 5, characterized in that, Steps (1) and (2) use the same Trichoderma harzianum solid inoculum.

7. The method according to claim 1, characterized in that, The rice mentioned is either conventional indica rice or conventional japonica rice.

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