Method for artificial breeding of cydia pomonella granulosis and its application in prevention and treatment of cydia pomonella

Abstract

The application discloses a method for artificially breeding Cotesia congregata and applying the Cotesia congregata to biological control of Malacosoma americanum, and determines optimal age, optimal parasitization ratio and optimal temperature of host Malacosoma americanum larvae for artificially breeding Cotesia congregata, effectively improves parasitization ratio and offspring quantity of the Cotesia congregata, shortens breeding cycle and improves bee breeding efficiency. Meanwhile, the optimal scheme for prolonging shelf life of the Cotesia congregata by storing cocoons or newly eclosed adults at low temperature is also determined. The new natural enemy product breeding application technology provided by the application is specially applied to biological control of Malacosoma americanum larvae, enriches existing Malacosoma americanum natural enemy product types, provides a more comprehensive solution for biological control of Malacosoma americanum, reduces overuse of chemical pesticides, and provides a powerful technical guarantee for balance and health of an ecological system.

Description

Technical Field

[0001] This invention relates to the field of biological pest control technology, and in particular to an artificial breeding method for the natural enemy insect, the American white moth, the *Gnaphalium affine*, and its application in controlling the American white moth. Background Technology

[0002] American white moth Hyphantriacunea The fall webworm (Erebidae), belonging to the order Lepidoptera, family Noctuidae, is a major forestry quarantine pest in my country. Native to North America, it invaded Dandong, Liaoning Province, my country in 1979, and now causes damage in 596 county-level administrative regions across 13 provinces. Biological control, as a crucial component of integrated pest management, relies heavily on the utilization of natural enemies. Statistics show that the fall webworm has 128 predatory natural enemies and 76 parasitic natural enemies in its native North America. This rich diversity of natural enemies is a key reason why the fall webworm has not caused significant damage in its native habitat. Since the late 20th century, my country has conducted detailed surveys and research on the natural enemies of the fall webworm, discovering abundant resources of natural enemies that can parasitize different life stages. Furthermore, the dominant natural enemy of the fall webworm during its pupal stage—*Triplophysa zhouensis*—has been successfully developed. Chouioiacunea This provides strong technical support for the green control of the fall webworm. However, the only natural enemy insect that can be used for the biological control of the fall webworm has been the parasitic wasp *Agrostis zhouensis*, and there has been a persistent shortage of natural enemy products targeting other life stages of the fall webworm. The fall webworm larvae feed extensively on plant leaves, which is the main period of damage and also the critical period for control. Due to the lack of natural enemy products for controlling the larvae, the control of the fall webworm during the larval stage still relies heavily on chemical pesticides, causing environmental pollution. Developing a natural enemy product that targets the fall webworm larvae is of great significance for constructing a green control technology system for the entire life cycle of the fall webworm, improving the integrated control effect, and maintaining ecological security.

[0003] White moths gather on a silken cocoon wasp Cotesia gregalis This is a parasitic wasp that infests the larvae of the fall webworm (Hemiberlesia spp.). Recent studies investigating the richness of the fall webworm's natural enemy groups have found that this wasp is the most dominant species in the natural enemy complex during the fall webworm's larval stage, exhibiting strong control over the fall webworm population. It is distributed in many areas of North and Northeast China, demonstrating strong adaptability. Currently, there is only literature describing this new species of *Brachystomum spp.*, and its application in the biological control of the fall webworm remains unknown. As a natural enemy insect with great potential, solving its artificial breeding techniques and identifying key aspects of its application are fundamental to its commercialization and application in forestry production. Based on this, we have innovated the breeding methods of *Brachystomum spp.* and its application in the control of the fall webworm. Summary of the Invention

[0004] The purpose of this invention is to provide a method for artificially breeding the white moth aggregation wasp, and a method for using artificially bred natural enemies to control the American white moth in forests.

[0005] Therefore, the present invention provides a method for artificially breeding the white moth aggregation disc wasp, the method comprising the following steps:

[0006] (1) Use the 4th to 7th instar larvae of the American white moth as hosts and raise them in petri dishes;

[0007] (2) The mated female parasitic wasps were introduced into a culture dish with a host and placed in an artificial climate chamber for cultivation. The ratio of parasitic wasps to the host American white moth was 1:5 to 5:1.

[0008] (3) After the parasitic behavior of the American white moth congregates on the velvet wasp, the American white moth larvae in the petri dish are kept in the same artificial climate chamber and fed regularly until the larvae mature, escape from the host and spin a cocoon or until they emerge as adults.

[0009] (4) Collect the cocoons of white moths and the adult bees that emerge from the cocoon of the parasitic wasp, and store them in an artificial climate chamber for use in the forest.

[0010] In a specific implementation, in step (1), the optimal age of the American white moth larvae used for bee breeding is 4 or 5 instars.

[0011] In a specific implementation, in step (1), in order to facilitate large-scale inoculation and breeding, a petri dish with a size of 90 mm × 15 mm is used, and the density of American white moth larvae is 25 per dish.

[0012] In a specific implementation, in step (2), the ratio of the number of white moth aggregation wasps to the number of host white moths is 3:5 to 1:1, preferably 3:5.

[0013] In a specific implementation, in step (2), the artificial climate chamber conditions are 20~30℃, RH=60%, photoperiod L:D=16:8 h, and the temperature is preferably 25℃.

[0014] In a specific implementation, in step (4), the age of the parasitic wasp cocoons is 1 to 5 days when they are stored, preferably 3 days; the artificial climate chamber conditions are a temperature of 4 to 12°C, a relative humidity of RH=70%, a photoperiod of L:D=16:8 h, a temperature of 10°C, and a storage time of no more than 30 days.

[0015] In the specific implementation, in step (4), the adult bee storage requires the collection of individuals that have just emerged from their molts; the artificial climate chamber conditions are a temperature of 5 to 25°C, a relative humidity of RH=70%, a photoperiod of L:D=16:8 h, a temperature preferably of 15°C, and a storage time not exceeding 25 days.

[0016] This invention also provides a method for using the white moth aggregation wasp obtained according to the above method to control the fall webworm in forests, the method comprising the following steps:

[0017] (1) After discovering the webs of the fall webworm in the forest, investigate the number of webs of fall webworm and estimate the number of larvae in the webs based on the size of the webs to obtain the population density of fall webworm in the forest.

[0018] (2) Based on the insect population density obtained from the survey, release adult wasps or cocoons of the white moth magpie in the forest, and investigate the parasitism of the white moth larvae by the white moth magpie magpie 7 to 10 days later to evaluate the biological control effect.

[0019] In a specific implementation, in step (1), a random sampling method is used to investigate the number of American white moth webs in the forest and the insect population density within the webs.

[0020] In a specific implementation, in step (2), the white moth gathering disc wasps need to be released in the early stage of the American white moth larvae breaking through the web; when releasing female adult wasps, the release ratio is 3:1 to 1:3, preferably 1:1; when releasing cocoons, the release ratio is 4:1 to 1:4, preferably 2:1.

[0021] This invention, by determining the parasitism preference of the parasitic wasp *Brachystomum glomeratum* for larvae of the fall webworm at different instars, clarifies the optimal age of the host for wasp breeding and the best control period for releasing the parasitic wasps. This improves the efficiency of artificial wasp breeding while enhancing the control effect in the field. Research on the optimal wasp-to-larva ratio revealed that the parasitism rate at a wasp-to-larva ratio of 3:5 was not significantly different from that at 4:5 or 5:5, but was significantly higher than that at 2:5 or 1:5. Furthermore, at a wasp-to-larva ratio of 4:5 or 5:5, no more parasitic wasp offspring were obtained compared to the 3:5 treatment, indicating that excessive input of female wasps did not significantly improve the reproductive efficiency of the natural enemy. These findings demonstrate that a high parasitism rate and a large number of offspring can be ensured while minimizing the number of female wasps required. Meanwhile, this invention also determined the optimal breeding conditions: at 25℃, RH=60%, and a photoperiod L:D=16:8 h, the parasitic wasp *Parasporium glomeratum* completes a generation in only 15 days, significantly shortening the breeding cycle and improving breeding efficiency. More importantly, the storage method for *Parasporium glomeratum* proposed in this invention significantly extends the shelf life of the natural enemy, facilitating a one-time flooding release of natural enemies during control. This invention also clarifies the optimal release period and optimal release ratio of *Parasporium glomeratum* when used in the field to control the fall webworm, maximizing the reduction of fall webworm damage while minimizing the use of natural enemies, thereby reducing the cost of biological control. This invention provides a new natural enemy product breeding and application technology specifically for biological control during the fall webworm larval stage. It not only enriches the existing variety of fall webworm natural enemy products but also provides a more comprehensive solution for biological control of the fall webworm throughout its entire life cycle, reducing the excessive use of chemical pesticides and providing strong technical support for the balance and health of the ecosystem. Attached Figure Description

[0022] Figure 1 American white moth larvae were reared in petri dishes.

[0023] Figure 2 White moths gather on discs where male and female adult wasps mate.

[0024] Figure 3 The white moth gathers on the disc of the silkworm and the silkworm wasp emerges from the body of the American white moth larva and spins a cocoon. Detailed Implementation

[0025] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings:

[0026] Example 1:

[0027] (1) Parallel control setup: Fall webworm larvae of different instars were selected as hosts for bee breeding, including treatment 1:3rd instar; treatment 2:4th instar; treatment 3:5th instar; treatment 4:6th instar; and treatment 5:7th instar. Five fall webworm larvae of the same instar were placed in a petri dish (90 mm × 15 mm). As an example, Figure 1 This demonstrates how to raise fall webworm larvae in a petri dish.

[0028] (2) One mated but parasitic female adult bee was introduced into each culture dish and placed in an artificial climate chamber at 25°C, photoperiod L:D = 16:8h, and RH = 60%. This was counted as one replicate, and each treatment was replicated 30 times. As an example, Figure 2 The image shows the mating of male and female adult wasps of the white moth congregating on a cluster of disc-shaped velvet wasps.

[0029] (3) Remove the female wasps 24 hours after receiving the wasps, and regularly change the artificial feed for the American white moth until the parasitic wasps develop to the point of escaping the host insect body, forming a cocoon and pupating.

[0030] (4) After the offspring wasps escape from the host insect and spin cocoons, the parasitism rate of the white moth's *Brachystomia solani* on hosts of different instars is counted. Successful parasitism is defined as the presence of parasitic wasps escaping from the host insect. As an example, Figure 3 It demonstrates how parasitic wasps burrow into cocoons and pupate after escaping from their bodies.

[0031] (5) The parasitism rate of female wasps on hosts of different ages, the number of parasitized wasps, and the developmental stages were statistically analyzed. The results are shown in Table 1:

[0032] Table 1. Parasitism rate, developmental duration, and number of offspring when *Coprinus micranthum* larvae are parasitized by *Baracus micranthum* sp. *Aegilops micranthum* larvae of different instars.

[0033]

[0034] As shown in Table 1, the parasitism rate of *Brachystomum simulans* on *Leymus chinensis* larvae of different instars differed significantly, showing that the parasitism rate first increased and then decreased with increasing host instar (F...). 4, 145 = 45.254, P<0.001). When the fall webworm parasitizes 4th or 5th instar larvae, the average parasitism rate can reach over 70%, but the average parasitism rate for 3rd instar larvae is only 15.33%. This phenomenon may be due to the difference in insect size. 3rd instar larvae of the fall webworm are relatively small, making it difficult for the parasitic wasp's ovipositor to accurately pierce them; however, 6th and 7th instar larvae are much larger and more resistant to parasitic wasp attacks, leading to a decrease in parasitism rate as the female wasp cannot fully subdue the host. Simultaneously, the host's instar also significantly affects the number of offspring (F1). 4, 123= 13.930, P<0.001). Using 4th or 5th instar fall webworm larvae as breeding hosts, introducing a single female wasp yielded more than 60 offspring wasps, making it the optimal scheme among all treatments. Furthermore, the host instar also had a certain impact on the developmental period of the parasitic wasps; after parasitizing 3rd or 4th instar hosts, the egg-larval stage of the parasitic wasps was significantly prolonged (F...). 4, 123 =201.078, P<0.001), after parasitizing a 3rd instar host, the cocoon stage of the parasitic wasp was also significantly prolonged (F =201.078, P<0.001). 4, 123 = 11.658, P<0.001). The above results indicate that using 4th and 5th instar larvae of the fall webworm as hosts, female wasps can exert the strongest parasitic effect and reproduce more parasitic wasp offspring; when the host is a 5th instar larva, the offspring wasps develop faster, indicating a shorter breeding cycle, suggesting that the 5th instar larvae of the fall webworm are most suitable for breeding the fall webworm parasitic wasp.

[0035] Example 2:

[0036] (1) Five fifth-instar larvae of the American white moth were introduced into a petri dish (90 mm × 15 mm) and artificial feed was provided.

[0037] (2) Parallel experimental setup: One mated female *Pteris vittata* wasp was inoculated into each culture dish and cultured in an artificial climate chamber under the following temperature conditions: Treatment 1: 15℃; Treatment 2: 20℃; Treatment 3: 25℃; Treatment 4: 30℃; Treatment 5: 35℃. Photoperiod L:D = 16:8 h, relative humidity RH = 60%. Each culture dish was counted as one replicate, and each temperature treatment was replicated 30 times.

[0038] (3) Remove the female wasps 24 hours after receiving the wasps, and regularly change the artificial feed for the American white moth until the parasitic wasps develop to the point of escaping the host insect body, forming a cocoon and pupating.

[0039] (4) After the offspring wasps escape from the host insect and spin cocoons, the parasitism rate of the white moth gathering disc wasps on the host at different ages is counted. Successful parasitism is defined as the presence of parasitic wasps escaping from the host insect.

[0040] (5) The parasitism rate, number of parasitized wasps, developmental period, and emergence rate of female wasps at different temperatures were statistically analyzed. The results are shown in Table 2:

[0041] Table 2. Parasitism rate, developmental period, number of offspring, and emergence rate of *Coprinus sphaeroides* at different temperatures.

[0042]

[0043] As shown in Table 2, the parasitism rate of *Coprinus macranthum* varies significantly under different temperatures. The parasitism rate initially increases and then decreases with rising temperature (F...). 4, 145= 33.210, P<0.001). Under treatments 2 and 3 (20℃ and 25℃), the parasitism rate was significantly higher than in other groups. Under treatment 1, the parasitic wasps showed lower activity and weaker parasitism. Under treatments 4 and 5, sustained high temperatures easily led to heat stress and death in female wasps, resulting in a decrease in the parasitism rate. Simultaneously, under both low and high temperature stress, the growth and development of offspring parasitic wasps were inhibited. Under the temperatures of treatments 2 and 3, parasitic wasp eggs and larvae could grow and develop better within the host, resulting in a significantly higher number of offspring under these conditions compared to other treatments (F<0.001). 4, 129 = 32.354, P<0.001). Furthermore, the developmental duration of parasitic wasps varied significantly under different temperatures; specifically, with increasing temperature, both the egg-larva and cocoon stages were significantly shortened (egg-larva stage: F...). 4, 129 =1373.399, P<0.001; Pupal stage: F 3, 104 = 2651.558, P<0.001). When the temperature is between 15 and 20℃, the development rate of the parasitic wasp slows down significantly, resulting in a significant extension of the reproductive cycle of the white moth aggregater wasp. At 35℃, the white moth aggregater wasp fails to emerge due to prolonged high temperature stress, while the emergence rate of the parasitic wasp is significantly higher at 20-30℃ than at 15℃ (F = 2651.558, P<0.001). 3, 96 = 355.524, P<0.001). In conclusion, at a temperature of 25℃, the parasitism rate of the parasitoid wasp *Pteris vittata* is high, the number of offspring is large, and the reproductive cycle is short, enabling the wasps to reproduce rapidly and in large numbers in a short period of time, which is the optimal temperature for wasp breeding.

[0044] Example 3:

[0045] (1) Inoculate 25 fifth-instar larvae of the American white moth into a petri dish (90 mm × 15 mm) and provide artificial feed.

[0046] (2) Parallel experimental setup: Each culture dish was inoculated with mated female *Paraceras fasciculata* at the following ratio (number of female wasps: number of hosts): treatment 1: 1:5; treatment 2: 2:5; treatment 3: 3:5; treatment 4: 4:5; and treatment 5: 5:5. After inoculation, the dishes were placed in an artificial climate chamber at 25℃, L:D = 16:8 h, and RH = 60%. Each culture dish was counted as one replicate, and each inoculation ratio treatment was replicated 10 times.

[0047] (3) After receiving the bees for 24 hours, the female bees are removed and the host continues to be fed artificial feed in the artificial climate chamber until the parasitic bees develop to the point of escaping the host body and pupating.

[0048] (4) After the offspring wasps escape from the host insect and spin cocoons, the parasitism rate of the white moth gathering disc wasps on the host at different ages is counted. Successful parasitism is defined as the presence of parasitic wasps escaping from the host insect.

[0049] (5) The parasitism rate, offspring number, and sex ratio of female bees under different inoculation ratios were statistically analyzed. The results are shown in Table 3:

[0050] Table 3. Parasitism rate, number of offspring, and percentage of females of the white moth *Gnaphalium affine* at different inoculation ratios.

[0051]

[0052] Table 3 shows that the parasitism rate of *Coprinus pachycarpa* varies significantly under different inoculation ratios. Specifically, the average parasitism rate gradually increases with the increase in the number of female wasps (F...). 4, 45 = 75.066, P<0.001). Under treatments 2, 3, and 4 (inoculation ratios of 1:5 to 3:5), the parasitism rate of parasitic wasps increased significantly with increasing wasp-to-parasite ratio, but the growth rate gradually slowed down under inoculation ratios of 3:5 to 5:5. This result indicates that when there are too many female wasps in a limited space, it may cause competition between female wasps and the host, which is not conducive to fully utilizing the parasitism ability of female wasps. At the same time, there were significant differences in the number of offspring of parasitic wasps under different inoculation ratios; the number of offspring in treatments 3 and 4 was significantly higher than in other treatments (F<0.001). 4, 45 = 283.148, P<0.001), and the number of offspring showed a trend of first increasing and then decreasing with the increase of the number of inoculated female wasps. Specifically, at an inoculation ratio of 5:5, although the parasitism rate of the parasitic wasps was high, the number of offspring was significantly lower than that at inoculation ratios of 3:5 to 4:5, indicating that the parasitic wasps failed to achieve optimal reproductive efficiency at high densities. Furthermore, different inoculation densities also significantly affected the proportion of female offspring; as the number of female wasps increased, the female ratio of the offspring gradually decreased (F...). 4, 45 = 63.427, P<0.001). Under inoculation ratios of 4:5 to 5:5, the proportion of female offspring was significantly lower than in other treatments. Overall, when the inoculation ratio of wasps was 3:5, the parasitism rate and the number of offspring were higher, and there was no significant difference compared with treatments 4 and 5 (inoculation ratios of 4:5 or 5:5), but the proportion of female parasitized wasp offspring was significantly higher in the latter. These results indicate that using an inoculation ratio of 3:5 for wasps in artificial bee breeding can save on the input of inoculated wasps while obtaining more female offspring, and is the optimal inoculation ratio for artificial bee breeding.

[0053] Example 4:

[0054] (1) Inoculate 15 fifth-instar larvae of the American white moth into a petri dish (90 mm × 15 mm) and provide artificial feed.

[0055] (2) Nine mated female parasitic wasps were introduced and placed in an artificial climate chamber at 25℃, L:D= 16:8 h, RH= 60% for cultivation until the parasitic wasps develop to the point of escaping the host insect and pupating in a cocoon.

[0056] (3) Parallel experimental setup: Three-day-old cocoons were collected in petri dishes (90 mm × 15 mm) and placed in the following artificial climate chambers: Treatment 1: 4℃; Treatment 2: 6℃; Treatment 3: 8℃; Treatment 4: 10℃; Treatment 5: 12℃. The photoperiod and relative humidity for each temperature treatment were L:D = 16:8 h and RH = 70%. Each treatment contained 30 cocoons and was counted as one replicate. Ten replicates were set up for each treatment.

[0057] (4) After 30 days of low-temperature storage, each treatment was transferred to an artificial climate chamber at 25℃, L:D = 16:8 h, and RH = 60%, and the eclosion success rate of cocoons stored at different temperatures was statistically analyzed. The results are shown in Table 4:

[0058] Table 4. Emergence rate of adult bees after cocoons are stored at different temperatures.

[0059]

[0060] Table 4 shows that the emergence rate of white moth cocoons and bee cocoons after storage under different low-temperature conditions varies significantly (F). 4, 45 = 86.720, P<0.001). The emergence rates of parasitic wasps stored under treatments 4 and 5 were significantly higher than those under other treatments, at 81.43% and 86.21%, respectively. This result indicates that storing parasitic wasp cocoons at 10–12°C is beneficial for extending the shelf life of *Pteris vittata*.

[0061] Example 5:

[0062] (1) Inoculate 15 fifth-instar larvae of the American white moth into a petri dish (90 mm × 15 mm) and provide artificial feed.

[0063] (2) Nine mated female parasitic wasps were introduced and placed in an artificial climate chamber at 25℃, L:D= 16:8 h, RH= 60% for cultivation until the parasitic wasps developed to adulthood.

[0064] (3) Parallel experimental setup: Newly emerged female bees were collected in plastic containers (8.5 cm in diameter and 8.5 cm in height) and placed in artificial climate chambers as follows: Treatment 1: 5℃; Treatment 2: 10℃; Treatment 3: 15℃; Treatment 4: 20℃; Treatment 5: 25℃. The photoperiod and relative humidity for each temperature treatment were L:D = 16:8 h and RH = 70%. Each treatment contained 100 female bees, and the survival status of the female bees was recorded daily until all female bees died. Each bee was counted as a replicate, and the average lifespan of the female bees under different storage conditions was statistically analyzed. The results are shown in Table 5:

[0065] Table 5. Lifespan of female bees under different storage temperatures

[0066]

[0067] Table 5 shows that when the storage conditions were 15℃, L:D = 16:8 h, and RH = 70%, the average lifespan of the white moth aggregation disc wasp was significantly higher than that of other treatments (F... 4, 495 = 964.242, P<0.001). At 5℃, the average lifespan of female wasps was significantly lower than at other temperatures, indicating that excessively low temperatures affect the lifespan of female wasps. This may be related to the reduced metabolic rate and slower physiological activity of parasitic wasps caused by low temperatures. These results indicate that 15℃ is the optimal storage temperature for preserving adult female *Amanita muscaria* wasps, and storing female wasps at this temperature is beneficial for preserving the wasp population and extending the shelf life of natural enemies.

[0068] Example 6:

[0069] (1) Based on the breeding methods and conditions determined in the previous experiment, white moth aggregation disc velvet wasps were bred.

[0070] (2) During the fourth instar larval stage of the American white moth, five independent forest plots with relatively consistent levels of damage caused by the American white moth were selected. The number of webs and the density of larvae of the American white moth in the forest were investigated by field survey.

[0071] (3) Parallel experimental setup: Based on the number of American white moth larvae in each sample plot, release female white moth aggregation wasps in the following ratios: treatment 1: wasp-to-larva ratio = 3:1; treatment 2: wasp-to-larva ratio = 2:1; treatment 3: wasp-to-larva ratio = 1:1; treatment 4: wasp-to-larva ratio = 1:2; treatment 5: wasp-to-larva ratio = 1:3.

[0072] (4) Seven days after the release of wasps, five trees were randomly selected from each plot of land that had been damaged. 250 fall webworm larvae were collected from each tree and brought indoors. They were reared in culture dishes (150mm × 15mm) with artificial feed at a density of 50 larvae / dish. The plants were cultured in an artificial climate chamber at 25℃, L:D = 16:8 h, and RH = 60%, until the parasitic wasps emerged from the host insects and pupated. Each tree was considered a replicate. The parasitism rate and the number of fall webworm larvae that could be bred from the parasitized fall webworms were statistically analyzed under different release ratios. The results are shown in Table 6.

[0073] Table 6. Control effect of different release ratios of female parasitic wasps on the white moth aggregation disc.

[0074]

[0075] Table 6 shows that the parasitism effect of *Conophytum commune* on the larvae of *Pteris vittata* varies significantly under different release ratios. Specifically, the parasitism rate and offspring number under treatments 1, 2, and 3 are significantly higher than those under other treatments (F...). 4, 20 = 24.868, P<0.001; F 4, 20 = 93.698, P<0.001). When the parasitic wasp ratio was 3:1 to 1:1, there was no significant difference in the average parasitism rate and the number of offspring. When the ratio was 1:2, the average parasitism rate and the number of offspring showed a significant downward trend. When the ratio was 1:3, the average parasitism rate dropped to 18.48%, and the number of offspring obtained was nearly twice that of the ratio of 3:1. In summary, treatment 3, i.e., releasing parasitic wasps with a ratio of 1:1, can maintain a high parasitism rate and the number of parasitic wasp offspring while saving the number of female wasps required. It can minimize control costs while meeting the requirements of control effectiveness and sustainable control strategies. Therefore, when releasing female parasitic wasps of the white moth *Papilionophora* in the forest to control the American white moth, a release ratio of 1:1 should be given priority.

[0076] Example 7:

[0077] (1) Based on the breeding methods and conditions determined in the previous experiment, white moths were bred and the cocoons that had not yet emerged were used as the target for release in the forest.

[0078] (2) During the fourth instar larval stage of the fall webworm, seven independent forest plots with relatively consistent fall webworm damage were selected. The number of fall webworm webs and larval density in the forest were investigated using the field survey method.

[0079] (3) Parallel experimental setup: Based on the number of American white moth larvae in each sample plot, release cocoons of white moth aggregation wasps according to the following wasp-insect ratios: treatment 1: wasp-insect ratio = 4:1; treatment 2: wasp-insect ratio = 3:1; treatment 3: wasp-insect ratio = 2:1; treatment 4: wasp-insect ratio = 1:1; treatment 5: wasp-insect ratio = 1:2; treatment 6: wasp-insect ratio = 1:3; treatment 7: wasp-insect ratio = 1:4.

[0080] (4) Ten days after the release of wasps, five trees were randomly selected from each plot of land that had been damaged. 250 fall webworm larvae were collected from each tree and brought back indoors. Artificial feed was provided in culture dishes (150mm × 15mm) at a density of 50 larvae / dish. The plants were cultured in an artificial climate chamber at 25℃, L:D = 16:8 h, and RH = 60%, until the parasitic wasps emerged from the host insects and pupated. Each tree was considered a replicate. The parasitism rate and the number of fall webworm larvae that could be bred from the parasitized fall webworms were statistically analyzed under different release ratios. The results are shown in Table 7.

[0081] Table 7. Control effects of different release ratios of white moth aggregation discs, silkworm cocoons, and bee cocoons.

[0082]

[0083] Table 7 shows that treatments 1, 2, and 3, with higher parasitism ratios (Fparasitism), had significantly higher parasitism rates and progeny wasps than other treatments (parasitism rate: Fparasitism). 6, 28 = 143.636, P<0.001; Number of offspring bees: F 6, 28 = 399.494, P<0.001). When the wasp-to-insect ratio was 2:1 to 4:1, there was no significant difference in the average parasitism rate and the number of progeny natural enemies. When the wasp-to-insect ratio was 1:1 to 1:4, the average parasitism rate and the number of progeny wasps showed a significant decreasing trend. When the wasp-to-insect ratio was 1:4, the parasitism rate decreased to 9.36%, and the number of progeny natural enemies was also reduced by nearly 5 times compared to treatment 1. In conclusion, when using the cocoons of the white moth-aggregating wasp as the release target, releasing natural enemies in the forest at a wasp-to-insect ratio of 2:1 can achieve a high control effect with relatively low cost, and is the optimal release ratio for releasing the cocoons of the white moth-aggregating wasp in the wild to control the fall webworm.

Claims

1. A method of controlling Malacosoma americanum with Cotesia rubecula, characterized by, The method comprises the following steps: (1) using 4th or 5th instar larvae of Malacosoma neustria as hosts, and feeding them in culture dishes at a density of 15-25 larvae per 90 mm x 15 mm culture dish; (2) introducing the mated C. aggregata female wasps into the culture dishes with hosts, and placing them in an artificial climate chamber for cultivation, wherein the number ratio of C. aggregata to the hosts M. neustria is 3:5; (3) after the parasitization of C. aggregata is completed, continuing to feed the M. neustria larvae in the culture dishes in the same artificial climate chamber, and adding feed regularly, until the larvae develop into mature larvae that escape from the hosts and form cocoons or until they emerge; (4) collecting the cocoons of C. aggregata or the emerged adult wasps, and placing them in an artificial climate chamber for storage, for application in forests; (5) after the M. neustria netting is found in forests, investigating the number of the netting, and estimating the number of the larvae in the netting according to the size of the netting, to obtain the population density of M. neustria in the forests; (6) releasing the adult wasps or cocoons of C. aggregata in the forests according to the population density obtained by the investigation, and investigating the parasitization of the M. neustria larvae by C. aggregata after 7-10 days, to evaluate the biological control effect. In the fourth step, the cocoon age of the parasitic wasps is 3 days when stored; the artificial climate chamber conditions are temperature 10℃, relative humidity RH=70%, light cycle L:D=16:8 h, and the storage time is not more than 30 days. In the fourth step, the emerged adult wasps are collected immediately after emergence; the artificial climate chamber conditions are temperature 15℃, relative humidity RH=70%, light cycle L:D=16:8 h, and the storage time is not more than 25 days. In the sixth step, the C. aggregata is released at the initial stage of the M. neustria larvae breaking the netting; when the female adult wasps are released, the ratio of the wasps to the hosts is 1:1; when the cocoons are released, the ratio of the wasps to the hosts is 2:

1.

2. The method for controlling the fall armyworm by using Cotesia flavipes according to claim 1, characterized in that, In the second step, the artificial climate chamber conditions are temperature 20-30℃, RH=60%, light cycle L:D=16:8 h.

3. The method for controlling the fall webworm using the parasitic wasp *Bombyx moth* according to claim 2, characterized in that... In the second step, the temperature of the artificial climate chamber is 25℃.

4. The method for controlling the fall webworm using the parasitic wasp *Bombyx moth* according to claim 1, characterized in that... In the fifth step, the random sampling method is used to investigate the number of the M. neustria netting in the forests and the population density of the hosts in the netting.

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